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Honours Projects 2012
Chemistry and bioChemistry
faCulty of life and physiCal sCienCes
Chemistry and Nanotechnology
Genetics and Biomedical Science
Forensic Science
Biochemistry and Molecular Biology
Biochemistry & Chemistry
2012 Honours
If you are interested in undertaking Honours at UWA, you may be already asking about the
exciting prospects available within each of the Disciplines and sub-disciplines comprising the
School. These include Biochemistry and Molecular Biology, Biomedical Science, Chemistry,
Forensic Chemistry, Nanotechnology, Genetics, and Structural Biology.
This Honours Project book and the associated School Honours Expo are intended to help you
explore the possibilities for 2012.
If you intend to enrol in Honours in 2012, this booklet will provide you with a comprehensive
overview of the interests of our research groups as well as outlining specific Honours projects
that are available.
The Honours Expo is designed to showcase the depth and diversity of research being
undertaken in the School and will enable you to discuss particular projects or even discuss the
design of new ones.
We hope that you will enjoy our Expo and that it will serve as a good introduction to the
range of Honours projects available in the School for next year.
Professor M Spackman
Head of School
Honours Co-ordinators
Biochemistry and Molecular Biology
Winthrop Professor Alice Vrielink
Phone: 6488 3162
Genetics
Winthrop Professor George Yeoh
Phone: 6488 2986
or
Winthrop Professor Lawrie Abraham
Phone: 6488 3041
Chemistry
Assoc Professor Sam Saunders
Phone: 6488 3153
Forensic Chemistry
Winthrop Professor John Watling
Phone: 6488 4488
Nanotechnology
Dr Robert Woodward
Phone: 6488 2751
Table of Contents
Research Expertise Table Page I - IV
Project Descriptions Page 1 - 66
How to Apply Page 68
Project Preference Form Page 69
i
Research Expertise Supervisor Research Area Discipline Page
Lawrie Abraham
Genetics
Biochemistry
Biomedical Science
Molecular Biology
Genetics,
Biochemistry &
Biomedical Science
1
Peter Arthur
Oxidative stress
Dystrophy
Aging
Muscle
Diabetes
Bioinformatics
Biochemistry 3
Paul Attwood
Enzyme structure and function
Enzyme kinetics
Protein phosphorylation
Histidine kinases
Histidine phosphorylation.
Biochemistry &
Chemistry 5
Murray Baker
Catalysis
Nanotechnology
Surface science
Biological chemistry/medicine
Polymer science
Molecular recognition, and sensors
Chemistry 7
Charlie Bond
Structural Biology
Protein Crystallography
Protein:protein interactions
Protein:nucleic acid interactions
Gene regulation
Biochemistry &
Chemistry 9
Bernard Callus Apoptosis
Cancer Signalling
Genetics &
Biochemistry 11
Reto Dorta Organometallic Chemistry
Catalysis Chemistry 13
Ela Eroglu
Nanotechnology
Microalgae
Wastewater treatment
Biofertilizer
Nanotechnology 15
ii
Gavin Flematti
Natural products
Analytical Chemistry
Separation Science
Chemistry 17
Simon Grabowsky
Crystallography
Electron Density
Computational Chemistry
Chemistry 18
Peter Hartmann Physiology and biochemistry of milk
synthesis Biochemistry 20
Dylan Jayatilaka Theoretical
Computational Chemistry Chemistry 22
George Koutsantonis
Organometallic Chemistry
Inorganic Synthesis
Molecular Electronics
Chemistry 24
Martha Ludwig
Molecular evolution
Molecular genetic
Molecular cell biology
Photosynthesis
Genetics,
Biochemistry &
Molecular Biology
26
Thomas Martin
Signalling
Protein Interaction
Bimolecular Fluorescence
Complementation, 14-3-3 proteins
Plant Histone Deacetylases
Plant nitrilases
Molecular Biology
Sugar sensing in plants
Nitrogen sensing in plants
Sugar metabolism
Nitrogen metabolism
Biochemistry 28
Allan McKinley
Environmental Chemistry
Physical Chemistry
Analytical Chemistry
Medicinal Chemistry
Chemistry 30
Harvey Millar
Biochemistry plant mitochondria
Oxidative stress and antioxidant
defence
Plant glutathione-S-transferases
Protein mass spectrometry
Proteome analysis
Biochemistry 32
iii
Matthew Piggott
Synthetic Organic Chemistry
Medicinal Chemistry
Chemical Biology
Chemistry 34
Colin Raston
Organic Synthesis
Tissue Engineering
Nano-chemistry
Graphene
Desalination Solar and Fuel Cell
Technology
Chemical Sensors
Drug Delivery
Microfluidics platforms
Nanotechnology 36
Sam Saunders
Atmospheric chemistry
Gas phase chemical kinetics
Reaction mechanisms
Computational chemistry
Chemistry 38
Ian Small
Genomics
RNA biology
Bioinformatics
Biochemistry 40
Steve Smith
Genomics
Genetics
Cell biology
Biochemistry
Bioinformatics
Systems biology
Metabolomics.
Biochemistry 42
Mark Spackman Crystallography
Theoretical chemistry Chemistry 44
Scott Stewart
Synthetic Organic Chemistry
Natural Product Synthesis
Palladium Catalysed Reactions
Domino Reactions
Chemistry 46
Keith Stubbs
Carbohydrates
Glycobiology
Synthesis
Inhibitors
Enzyme kinetics
Chemistry 48
Swaminatha Iyer BioNanoChemistry Nanotechnology 50
iv
Robert Tuckey
Enzymes
Cytochrome P450
Vitamin D
Steroids
Hydroxylases, placenta
Skin cancer
Metabolism
Biochemistry 52
Daniela Ulgiati
Genetics
Biochemistry
Biomedical Science
Molecular Biology
Molecular Biology,
Biochemistry,
Genetics
54
Alice Vrielink
Protein structure
Crystallography
Enzyme mechanism
Structure-Function Relationships
Rationale Drug Design
Biochemistry &
Chemistry 56
John Watling Forensic Chemistry Forensic Science 58
Jim Whelan
Mitochondrial biogenesis
Gene regulation
Phosphate metabolism
Molecular cell biology
Genetics
Biochemistry,
Genetics, &
Biomedical Science
60
Duncan Wild
Physical Chemistry
Laser Spectroscopy
Mass Spectrometry
Van der Waals clusters
ab initio calculations
Chemistry 62
Michael Wise
Bioinformatics
Microbial informatics
Low complexity/natively unfolded
proteins
Computational evolutionary biology
Biochemistry 64
George Yeoh
Liver stem cell
Cancer
Cell therapy
Genetics,
Biochemistry 66
1
WINTHROP PROFESSOR
LAWRIE ABRAHAM Room 2.58, Bayliss building, Phone: 6488 3041,
Email: [email protected]
Human Molecular Biology Lab
Our group is interested in the transcriptional regulation of gene expression. We are also interested in the effects
of genetic polymorphism (SNPs) on the expression of genes, particularly promoter and other regulatory
variants.The focus is on genes that are involved in regulating inflammatory responses and understanding how
genetically determined differences in expression contribute to diseases such as autoimmune disease, cancer and
cardiovascular disease. To this end we are involved in the identification of transcription factors and upstream
components of the signal transduction pathways that regulate these genes. Our long-term aim is to develop
therapeutic strategies to modulate the activity of these genes through interference with such regulators in order
to prevent disease. Students will be exposed to a range of techniques including DNA sequencing, DNA cloning,
cell culture, transfection assays, RT-PCR, expression array analysis, siRNA knockdown, DNA binding assays
(EMSA), protein analysis, DNase I Footprinting, Chromatin immunoprecipitation (ChIP) and FACS analysis.
PROJECTS
1. The Transcriptional control of the CD30 Gene in Anaplastic Large Cell Lymphoma (Genetics,
Biochemistry or Biomedical Science)
Anaplastic large cell lymphoma (ALCL) is a variant of immunoblastic lymphoma and tends to be clinically
aggressive, resulting in the destruction of the involved lymph node structure, the infiltration of the lymph node
sinuses by large transformed neoplastic cells with prominent
nucleoli. The major diagnostic marker of ALCL is strong
overexpression of the CD30 gene thought to result from a
transforming event that leads to neoplasia. Fundamental to our
understanding of the causes and treatment of ALCL is an
understanding of the mechanism of overexpression of CD30.
The CD30 gene promoter, including an ALCL-specific
hypersensitive site we have discovered in the 1st intron, will be
characterised with respect to transcriptional control elements
by EMSAs, CD30 reporter gene analysis and CHART
(chromatin accessibility by real-time PCR). The transcription
factors binding to the promoter and the 1st intron will be
identified by use of a 2-dimensional proteomics technique
developed in our group. Once cloned, the identified proteins
will be tested for the ability to repress endogenous expression
and reporter constructs by overexpression in cell lines and by
RNAi approaches. Chromatin immunoprecipitation (ChIP) assays will also be carried out to establish the in vivo
relationship between the various cis-elements and trans-acting factors, including sites of histone modification. The
long-term aim is to develop therapeutic strategies that interfere with the transcriptional regulation of CD30 and so
block the deleterious effects resulting from overexpression of CD30.
2. Characterisation of functional variants of Vanin 1, a QTL controlling HDL-C Levels (Genetics,
Biochemistry or Biomedical Science)
This collaborative project with the Texas Biomedical Research Institute, USA involves the characterisation of the
Vanin 1 gene, which has been shown to be genetically associated with low levels of High Density Lipoprotein-
cholesterol ("good" cholesterol) levels in the blood. Low HDL levels are a strong risk factor for cardiovascular
diseases such as arthrosclerosis and heart attack. Twelve non-coding variants in the Vanin 1 gene were found that
fall into 4 isocorrelated redundant variant sets (IRVS) show significant correlations with HDL-C as well as Vanin
1 mRNA expression levels. The most likely functional promoter variant at -137 exhibits a strong association with
2
HDL-C levels (p = 0.002). The project aims are to
characterise transcription factors that differentially bind to
the IVRS variants using EMSA (see Fig) followed by
peptide mass fingerprinting and also to determine the
effects of the candidate functional SNPs on transcriptional
activity using reporter gene analysis. A further aim is to
identify modulators of VNN1 expression & determine their
effects on allele-specific transcription of VNN1 using
mRNA expression profiling. An understanding of how the
gene is controlled will inform the development of
therapeutic strategies and/or drugs to modulate the activity
of the Vanin 1 gene with the objective of raising HDL-
cholesterol levels in individuals at risk.
3. Mechanism of Action of Newly Synthesised Thalidomide Derivatives. (Biochemistry or Biomedical
Science)
Thalidomide is a synthetic glutamic acid derivative used in the 1950‘s as a treatment for insomnia and as an
antiemetic agent. Later investigations found that thalidomide had teratogenic properties. In a collaborative project
with Dr Scott Stewart, newly synthesised and potentially safe thalidomide-based drugs will be screened for novel
biological activities using TNF reporter gene assays. For those students interested in the functional aspects of
thalidomide and the newly synthesised derivatives, transcriptional profiling will be carried out, using Affymetrix
microarrays to define novel cellular activities, with a focus on therapeutic application. The project also involves
the identification of the cellular targets of thalidomide which will be informative in a more rational drug design.
Photoactivatible biotin-derivatized thalidomide will be used to treat cells, followed by UV-catalysed cross-linking
(see Fig). Proteins will be isolated and identified by biotin-streptavidin affinity chromatography and mass
spectrometry. The proteins identified will be validated with respect to their interaction with thalidomide and by
assessing functional aspects of the candidate proteins. Interactions will also be validated using confocal cell
imaging.
4. Identification of Genetic Variation in Preeclampsia by Whole-Genome Exome Sequencing (Genetics
or Biomedical Science)
The genetic analysis of preeclampsia continues to be one of the most critically important and unresolved areas
of obstetric medicine. There is currently no known cure for preeclampsia other than delivery of the baby. Like
many other common human diseases there is a large genetic component underlying susceptibility to developing
preeclampsia but the genetics are complex and not yet fully understood. This project is a collaboration with
W/Prof Eric Moses and involves the identification of functional genetic variants associated with preeclampsia.
The emphasis is on whole-genome exome sequencing in families and represents the current state-of-the-science
for genetic dissection of complex traits. The goal is to identify the specific genetic polymorphisms responsible
for susceptibility to preeclampsia with the view to informing the development of much-needed diagnostic
reagents and therapeutic strategies. This approach has been made possible by recent technological advances and
efficiencies in high-throughput next generation DNA sequencing. This project involves a multidisciplinary team
of investigators who have led the field in the recruitment and genetic analysis of preeclampsia and
cardiovascular disease in families. The collection of 72 preeclampsia families from Australia/New Zealand,
Finland, Iceland and Norway are the best available worldwide, making this a time of unprecedented opportunity
for finding the most likely functional variants influencing susceptibility to preeclampsia.
3
9BASSOCIATE PROFESSOR
PETER ARTHUR 10B
Room 2.41, Bayliss Building, Phone: 6488 1750
Email: [email protected]
Reactive Oxygen Species as modulators of signal transduction pathways and
biochemical systems
Oxidative stress is caused by reactive oxygen species (ROS) and is thought to exacerbate pathology associated
with many chronic diseases and conditions. Examples include Alzheimer‘s disease, atherosclerosis, dementia,
diabetes, emphysema, heart disease, HIV/AIDS, kidney disease, liver disease, muscular dystrophy, Parkinson's
disease, Rheumatoid arthritis, some cancers and aging. However, preventing the harmful effects of oxidative
stress is not a simple matter, as antioxidant treatments have generally been ineffective in the treatment of these
conditions.
One challenge has been the lack of understanding of the various molecular mechanisms by which oxidative
stress causes pathology. We have established that cysteine residues on proteins are particularly sensitive to
oxidative stress and our laboratory is playing a leading role in identifying proteins sensitive to oxidative stress.
Our work, and the work of others, has established that multiple proteins are sensitive to oxidative stress, which
means oxidative stress could have a widespread impact on many cellular processes (metabolic pathways, ion
transport, protein synthesis, protein degradation, gene expression, signal transduction pathways). Our work into
how oxidative stress affects cellular processes will offer new opportunities to treat oxidative stress.
This research area is constantly developing, so I am happy to discuss the research area in general or work with
you to develop a project that suit your interests. I am an experienced supervisor with a preference for
collaborative projects so that you can gain the benefits of dual supervision. Please see below examples of
current research projects to give you an idea of the type of work we do.
PROJECTS
1. How does oxidative stress affect cell signaling pathways?
Collaborative with Dr Thea Shavlakadze & Prof. Miranda Grounds, School of Anatomy and Human Biology
Insulin growth factor 1 (IGF-1) is a potential therapeutic agent for muscle ageing and muscular dystrophy. In
both conditions oxidative stress plays a significant damaging role, and has the potential to block the actions of
IGF-1. The objective of this project is to use a cell culture model to examine the effect of ROS on the function
of signal transduction proteins. This project will involve using proteomic technology including protein
separation techniques (HPLC, 2D gel electrophoresis, antibody technology) and protein identification
techniques (mass spectrometry). Additional techniques may include Immunohistochemistry, Western Blotting,
quantitative PCR and EMSA.
2. Does oxidative stress cause muscle wasting?
Collaborative with Dr Thea Shavlakadze & Prof. Miranda Grounds, School of Anatomy and Human Biology
As skeletal muscle ages it loses strength and power leading to reduced mobility and deleterious changes in
lifestyle. The relentless loss of muscle mass and function in elderly individuals impairs daily functions such as
walking, using stairs and rising from chairs and results in an increased incidence of falls. Muscle wasting is also
associated with immobility and diverse pathologies such as cancer, bacterial sepsis, AIDS, diabetes, and end-
stage heart, kidney, and chronic obstructive pulmonary disease. We are using transgenic mouse models of
muscular dystrophy (which we already have) and ageing (which we are developing) to investigate the role of
oxidative stress in muscle wasting. Transgenic mouse models are particularly significant in biomedical research
because they reflect the complexity of human disease processes.
The objective of this project is to establish whether oxidative stress causes changes in protein turnover in
muscle, since decreased protein will lead to muscle wasting. For this work a muscle cell line (C2C12) will be
used, as cell culture systems are particularly useful experimental systems to pin point the precise molecular
mechanisms involved in disease processes. Techniques likely to be required for this project include proteomic
techniques, tissue culture, quantitative PCR for atrophy related genes and measurement of oxidative stress. This
4
project is also related to our larger effort to understand the
effects of mild oxidative stress (particularly ageing) by
developing a transgenic mouse over-expressing catalase.
3. Oxidative stress in ageing mice
Collaborative with Dr Thea Shavlakadze & Prof. Miranda
Grounds, School of Anatomy and Human Biology
The trend of ageing populations in many countries has become a
significant concern because age itself is a key risk factor for
many chronic degenerative diseases. Examples include
sarcopenia and neurodegenerative diseases such as Alzheimer's
and Parkinson's disease. Many of the age-dependent pathologies
been linked to oxidative stress, so targeted interventions aimed
at treatment or prevention of oxidative stress have the potential
to alleviate ageing pathologies. In this context, it is interesting
to note that both cardiac pathologies and cataract formation were
delayed in mitochondrial catalase knock-in mice.
One strategy for addressing the challenges posed by ageing
populations is to understand the molecular mechanisms
underlying the ageing process. We have hypothesized that oxidative stress affects susceptible proteins which
disrupt cellular homeostatic mechanisms and lead to pathological consequences including increased oxidative
stress sufficient to cause irreversible damage to cellular macromolecules. To develop evidence for these
hypotheses, a range of markers of oxidative stress will be used to assess how oxidative stress develops in ageing
mice. One experimental approach will involve using proteomics to identify proteins susceptible to developing
oxidative stress. A second experimental approach will test whether peroxiredoxins can be used as sensitive
indicators of oxidative stress. Peroxiredoxins are thought to be significant contributors to cellular removal of
hydrogen peroxide, yet are readily inactivated by oxidation of susceptible thiol groups. Inactivation of
peroxiredoxins may also have significant biological consequences by exacerbating oxidative stress.
4. Oxidative stress in Diabetes
Collaborative with Prof. Paul Fournier, School of Sport Science, Exercise and Health
Type 2 diabetes mellitus (T2DM) is a complex disorder that has reached epidemic proportions in Australia,
affecting nearly one sixth of its adult population above 40 years old. This is a source of much concern because
intensive personal and medical attention is required to manage and treat this condition. In addition, there is the
large burden of the many long-term microvascular and macrovascular complications associated with diabetes.
These include diabetic retinopathy which may lead to blindness, diabetic neuropathy with associated increased
risk of amputation and early death, diabetic nephropathy leading to end-stage renal disease, and macrovascular
complications such as stroke, coronary artery disease, and myocardial infarction.
Diabetes is characterised by impaired insulin secretion and a marked resistance to the action of insulin,
particularly in skeletal muscles. We have hypothesized that protein thiol oxidation is contributing to insulin
resistance. The objective this project is to establish whether oxidative stress is interfering in the function of key
proteins involved in the insulin signaling pathway (eg IRS1, AKT) in an animal model of insulin resistance
(high fat fed rats). This project will involve using proteomic technology including protein separation techniques
(HPLC, 2D gel electrophoresis, antibody technology) and protein identification techniques (mass spectrometry).
The effect of oxidative stress on proteins we will be evaluated using a patented technique developed by Dr.
Arthur.
5. Systems Approaches to Oxidative Stress
Collaborative with Professor Michael Wise
The objective of this project is to develop and use bioinformatic methods to identify the cellular processes and
organelles that are particularly sensitive to oxidative stress. This will involve categorizing the involvement of
proteins (those identified as sensitive to oxidative stress) in different cellular processes. You will be using
pathway analysis software such as IPA (www.ingenuity.com), keyword clustering software (Protein Annotators
Assistant) and databases such as BioCyc, Reactome and Kegg to look for common themes/processes. Protein-
protein interaction data and data about predicted location may also be useful.
Mdx/IG
F-1-1
Md
x
Figure 1. One year old male mdx/IGF-1
and mdx littermate mice. Mdx/IGF-1
transgenic mice are much bigger
compared to their age matched
littermates and they have a pronounced
skeletal muscle hypertrophy.
5
PROFESSOR PAUL ATTWOOD Room 3.69, Bayliss Building, Phone: 6488 3329
Email: [email protected]
The research focus of Prof. Attwood's laboratory is the structure and function of enzymes in general. However,
there is a particular focus on two enzymes:
1. Pyruvate carboxylase is a key biotin-dependent enzyme that provides oxaloacetate for the TCA cycle,
gluconeogenesis and neurotransmitter synthesis, whose structure we have just determined. There is also a
strong correlation between the activity of this enzyme and insulin secretion and thus an association with
Type II diabetes. We have determined the first structure of a biotin-dependent carboxylase holoenzyme, the
pyruvate carboxylase from Rhizobium etli:
St. Maurice et al. (2007) Science 317, 1076-1079.
We are investigating the structure-function relationships in this enzyme, with a combination of site-directed
mutagenesis, kinetic and physical methodologies. The ultimate aims of this project are to understand the
mechanism of action of the enzyme and design drugs that will act either as inhibitors (anti-fungals and anti-
bacterials) or stimulate the activity of the enzyme (diabetes treatment). We are currently working on the
mechanism of allosteric regulation of the enzyme by acetyl CoA and the mechanism of catalysis with respect
to half-of-the sites reactivity in the enzymic tetramer.
Suitable for students with a biochemistry or biochemistry/chemistry background.
6
2. Mammalian histidine kinases catalyse the phosphorylation of histidine residues in substrate proteins.
This is a little understood form of phosphorylation in mammalian cells and its biological roles are not yet
clear, although we have established a link between enhanced histone H4 histidine kinase activity and
hepatocellular carcinoma in human liver and shown it to be a possible oncodevelopmental marker of
hepatocellular carcinoma (see below). This discovery offers a potential target for treatment or diagnosis of
liver cancer.
HCCT = HEPATOCELLULAR CARCINOMA TISSUE
HCCN = NORMAL TISSUE SURROUNDING HEPATOCELLULAR CARCINOMA
NORMAL = NORMAL ADULT LIVER
Tan et al. (2004) Carcinogenesis 25, 1-6.
However, we really need to know more about the cellular role of histidine phosphorylation in general and
particlularly in histone H4. One of the difficulties in the investigation of histidine phosphorylation is the
detection of proteins containing phosphohistidine in cells and tissues, partly due to the lability of the P-N
bond and also because there are two isomers of phosphohistidine N1 and N3 (se below). To address this
problem I am currently collaborating with Dr. Matthew Piggott to develop pan-phosphohistone antibodies
for the detection of histidine-phosphorylated proteins, by synthesizing and using non-hydrolysable analogues
of phosphohistidine as immunogenic haptens (see triazole analogues below). This would be part of the
Honours project which would be jointly supervised by Prof. Attwood and Dr. Piggott and the chemistry
content could be adjusted to suit either a Chemistry major student or a biochemistry major student. Other
components could include some purification and characterization of histidine kinases.
HN
NH
O
proteinprotein
NN
P
O
O
O
1
3
HN
NH
O
proteinprotein
HNN 1
3
histidinekinase
ATP
histidine residue N1-phosphohistidineresidue
H3N
N
O
O
NN
H2N
N
O
O
NN
POO
OPO
OO
13
1
3
HN
NH
O
proteinprotein
N
N3-phosphohistidineresidue
N
POO
O 3
OR 1
stable triazole analogue stable triazole analogue
Suitable for students with a biochemistry or chemistry background.
7
PROFESSOR MURRAY BAKER Room 4.09, Bayliss building, Phone: 6488 2576
Email: [email protected]
My group's research interests are primarily in synthetic chemistry—we aim to apply our skills in synthesis to
problems in areas such as catalysis, nanotechnology, surface science, biological chemistry/medicine, polymer
science, molecular recognition, and sensors. Honours projects are currently available in the following areas.
1. Biodegradable and biocompatible materials for tissue engineering
Collaboration with Prof Traian Chirila (Prevent Blindness Foundation, Queensland), Prof Kathy Luo (Nanyang
Technological University (Singapore), Dr Keith Stubbs (UWA), and Dr David Brown (Curtin).
Biocompatible materials are materials that can be placed in contact with biological tissue without causing
infection or other undesirable biological responses. One of the most important biocompatible polymers is
poly(hydroxyethyl methacrylate) (PHEMA). PHEMA-based materials are made by co-polymerising
hydroxyethyl methacrylate (HEMA) with suitable crosslinking agents. PHEMA is
already used to fabricate permanent medical implants, such as the artificial cornea
developed by Prof Traian Chirila. An important feature of PHEMA is its ability to be
easily fabricated in a porous form that is suitable for hosting cell growth. The pictures
here show a scanning electron microscopy image of a sample of porous PHEMA (left)
and an optical microscope
image of a similar sample of
PHEMA after implantation
into a mouse (right). In the
latter image, blood vessels
and regenerating tissue
growing into the pores in the
PHEMA are clearly visible.
In collaboration with Prof Chirila, we are now developing biodegradable forms of PHEMA, for new
applications in tissue engineering. This research includes study of: (1) new biodegradable crosslinking agents
based on peptides and (with Dr Stubbs) carbohydrates); (2) new methods of controlled polymerisation of
HEMA; (3) incorporation cell adhesion factors and cell-growth factors into PHEMA; and (4) new forms of
biodegradable PHEMA (e.g., powders and thin films on surfaces) as substrates for tissue growth in the
laboratory.
In collaboration with Prof Luo we are investigating PHEMA as a component of polymer-cell conjugates to build
artificial tumours for use in cancer research. This work includes: (1) development of polymerisation methods
using non-toxic reagents and catalysts and (2) polymerisation of HEMA and HEMA oligomers in the presence
of live cells.
2. Chemical and Biological Applications of N-Heterocyclic Carbene Complexes
N-Heterocyclic carbenes (NHCs) are analogues of phosphines, but they
have some significant advantages, including ease of synthesis and strong
donor ability. NHC complexes are easily accessible via azolium ions (eg
1). We are exploring the synthesis and properties of interesting azolium
ions and transition metal NHC complexes. We have found that
complexes such as 2 are excellent catalysts for certain C-C bond forming
reactions. The unique ruthenium complex 3 is of interest as a potential
anti-cancer agent, since it is an analogue of a well-known class of anti-
cancer compounds such as 4.
Cationic Au(I) NHC complexes such as 5 and 6 exhibit activity
against certain cancer cell lines. This activity appears to be a consequence
of Au binding to an enzyme in mitochondria, and the selectivity for
N
N
N
N PdBr
Br
1
N
N
N
N
2
3
N
N
N
NCl
Ru
+
ClRu
+R
H2N
NH2
4
O
OOH
hydroxyethyl methacrylate
8
killing cancer cells over normal cells can easily be tuned by variation of the hydrophilic-hydrophobic character
of the NHC ligands.
N NN
N
N
N
Au
Au
2
NN
6
N N
Au
NN
Cl
Cl
N N
Au
NN
75
The Au(I)-NHC complexes are easy to synthesize and they offer the prospect of fewer toxic side-effects than
their better-known Au(I)-phosphine counterparts. Au(I) complexes such as 5 and 6 and Au(III) complexes such
as 7 also have exciting prospects as robust catalysts for a range of oxidation and C-C bond forming reactions.
An exciting goal in the Au-NHC area is to replace one of the NHC ligands with other ligands that have their
own innate biological activity. Thus, complexes of form [(NHC)-Au-L]+
have two potential modes of action:
deactivation of an enzyme by Au, and separate anti-cancer activity exhibited by L.
Another area of interest is the use of NHC-metal complexes as antibacterial agents to treat drug-resistant
infections. We have found that Au-NHC complexes such as 6 tend to concentrate in lysosomes of some cells,
and one way in which some bacteria resist drug treatments is by sequestering themselves in lysosomes. Thus,
any Au-NHC complexes (or Ag-NHC complexes, which are structurally similar to the Au complexes) that show
anti-bacterial activity have the potential to be therapeutic agents for some bacterial infections that are difficult to
treat using existing drugs.
3. Azamacrocycles and Catalysis of Organic Reactions by Iron Compounds
Triazacyclononanes (TACNs) are excellent ligands for transition metals. Numerous TACN complexes are
known, many have demonstrated interesting catalytic and biological activity, and some have served as model
systems for the active sites in metalloenzymes. The main disadvantage of TACN ligands is that their syntheses
are long and tedious.
Triazacyclohexanes (TACHs) are smaller analogues of TACN.
The chemistry of TACHs is relatively undeveloped, but TACHs
are easy to make (just one step from formaldehyde and a primary
amine) and they form many compounds analogous to TACN
complexes. Because the TACH ring is small, however, bonding
in TACH-metal complexes is much more strained than in
TACN-metal complexes, and so TACH complexes are quite
labile.
Recently, aminodiazacycloheptanes (ADACHs) have been proposed as analogues of TACN. ADACHs may be a
"happy medium" between TACHs and TACNs, since ADACHs are easy to synthesize and they offer a
coordination geometry similar to that of TACN. We have already used complexes such as the molybdenum
tricarbonyl adducts shown above to compare the chemistry of TACH, TACN, and ADACH systems.
Now we are starting a new project in this area, to examine new classes of iron complexes as potential catalysts.
Iron is the most abundant transition metal, it is very cheap, and it is non-toxic. Over the last few years, iron
compounds ranging from ferric nitrate through to tetrahedral iron phosphine complexes have been found to
catalyse organic transformations that previously had been achieved only by catalysts based on much more
expensive metals such as palladium. Catalysis by iron complexes is still in its infancy and is not well
understood, and selectivity is still poor in most cases. There are great opportunities for the development of
useful, cheap, and non-toxic catalysts based on iron. One way to address the problem of
selectivity in iron-catalysed reactions may be to bind the iron in a favourable coordination
environment, such as the environments provided by the facially-coordinating TACH and
ADACH ligands. These environments would bind the iron and so inhibit certain
unfavourable processes (eg formation of iron oxides) but leave three mutually cis
coordination sites for catalytic reactions to occur. A few Fe-TACH compounds are known.
The Fe(III) complex 8 would serve as a convenient starting point for this study.
N
N
N
Fe
ClCl
Cl
8
N N
NH2
Mo
C
N NH2
C
N
C
N NN
Mo
CC
C
N N
N
TACN
N
N
N
TACH
N
N
N
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9
PROFESSOR CHARLIE BOND 23BRoom 4.32, Bayliss Building, Phone: 6488 4406
Email: [email protected] U
Structural Biology
Structural Biology research involves building a three-dimensional picture of biological molecules to shed light
on the molecular interactions and events which drive many of the fundamental processes of life. Investigations
in my lab address proteins of relevance to human health, including nucleic acid processing proteins involved in
regulating gene expression, and enzymes essential to the survival of life-threatening parasites, which may be
drug targets.
Different aspects of this research can be tailored to students with strengths in Biochemistry, Chemistry, and
Biophysics. Structural Biology research typically involves the opportunity to learn from a diverse set of useful
techniques including molecular biology, protein purification and crystallisation, spectroscopy, X-ray
crystallography, molecular modelling, bioinformatics, unix computing. The Structural Biology lab is equipped
with state-of-the-art equipment including a crystallization robot and X-ray data collection facilities.
For further information, reprints of papers or to find out about other research in the lab come and see me (MCS
Rm 4.32) and look at HUhttp://www.crystal.uwa.edu.au/px/charlie UH .
13BPROJECTS
(The exact scope of each project will vary depending on the interests and experience of the student).
1. Structure of the paraspeckle interactome (suitable for more than one student)
Collaborative with Dr Archa Fox and DrSven Hennig (WAIMR)
An emerging and exciting research area is the role of noncoding RNAs in controlling gene expression.
‗Noncoding‘ RNAs are molecules that are functional as RNAs, and do not encode for proteins. Paraspeckles are
the first sub-nuclear structure known to form around a long noncoding RNA (lncRNA), making them an
important model system within lnRNA research. This is particularly relevant when it comes to cancer, as several
lncRNA have been shown to act as molecular scaffolds, recruiting proteins to form oncogenic complexes that
drastically alter gene expression leading to metastasis and ultimately poorer outcome for patients.
Paraspeckles contain a number of different proteins that
are either (1) responsible for paraspeckle formation (2)
required for paraspeckle function, or (3) are regulated by
sequestration within paraspeckles. The Bond lab has
recently solved the 3D structure of a number of homo- and
heterodimers of paraspeckle proteins (see figure 2). In an
effort to determine the roles of the other known
paraspeckle proteins in paraspeckle formation and
function, we are undertaking a large-scale interactome
analysis of paraspeckle components.
This project involves investigating interactions of key
paraspeckle proteins PSP2 and Matrin3 with other
paraspeckle proteins. It will involve mapping the domains
in each protein responsible for protein:protein interactions.
A number of techniques will be applied, including
molecular biology, yeast-two-hybrid assays, protein
expression in bacteria, purification and in vitro interaction
assays. The ultimate goal is to crystallise and solve the
structure of protein complexes. In many cases
sophisticated expression strategies are used such as co-expression of interacting proteins, in an effort to stabilise
interaction partners, leading to large-scale protein production. This project will provide important building
blocks for understanding how nuclear proteins together build up a lncRNA-structure, and how their
sequestration affects function.
Figure 1. Intermolecular interactions in paraspeckles
10
2. Structural studies of DBHS proteins – Key factors in gene regulation
Collaborative with Dr Mihwa Lee
Key paraspeckle proteins include the „Drosophila behavior/Human Splicing‟ (DBHS) family of proteins have
been implicated as important regulators of gene regulation in mammals. Here they are involved in a regulation
mechanism whereby mRNA molecules containing a particular structural motif are stockpiled in the nucleus so
they cannot be translated into protein. On a specific signal, the stockpiled mRNA is released and a burst of
protein production takes place. In mammals this process is controlled by three highly-conserved related DBHS
proteins which can form heterodimers which then form into large nuclear bodies called paraspeckles. The Bond
lab has recently solved the 3D structure of a number of homo- and
heterodimers of DBHS proteins (see Figure 2).
This project will build on this exising structural knowledge of DBHS
proteins to investigate the propensity of DBHS to for larger aggregates via
a coiled-coil interaction motif. Protein samples will be cloned, expressed in
bacteria, purified and studied by a panel of biophysical techniques
including crystallization and X-ray diffraction, dynamic light scattering and
analytical ultracentrifugation.
The student will learn the principles of basic molecular biology, protein
expression and purification, X-ray crystallography and complementary
biophysical techniques.
Figure 2. Crystal structure of a
DBHS protein heterodimer
from our lab
11
DR BERNARD CALLUS Senior Research Fellow
Room 3.49, Bayliss building, Phone: 6488 1107
Email: [email protected]
Apoptosis and Cancer Signalling
Our research group focuses on the mechanisms of apoptosis (programmed cell death) as well as the signalling
pathways that regulate cell death pathways. Particular focus is given to how the abnormal regulation of these
signalling pathways can contribute to the development of cancer. Typically, cancer cells are profoundly resistant
to apoptotic stimuli, e.g. chemotherapeutic drugs, radiation, and this apoptotic resistance is considered to be an
essential component in the development of tumours. Often this is due to amplification of oncogenes, e.g. Bcl-2,
or the loss of tumour suppressors, e.g. p53, or a combination of both which impart apoptotic resistance in cells.
Our research incorporates molecular biology and cellular based assays to examine the impact of increased
oncogene expression or loss of tumour suppressor gene expression in cells to examine how they impact on
apoptotic mechanisms as well as the signalling pathways that are regulated by them that ultimately contribute to
the development of cancer. Our research aims to identify novel regulators involved in apoptosis and cancer as
candidates for rational drug design leading to the development of new therapies to kill cancer cells.
Feel free to come and discuss your research interests and to learn more about our ongoing projects in the lab.
PROJECTS
1. The role of the Arf/Ink4a tumour suppressors on the proliferation, differentiation and transformation
of liver-progenitor cells. With Professor George Yeoh, Biochemistry and Molecular Biology.
We have previously shown that the expression of the Arf tumour suppressor is significantly down-regulated to
undetectable levels during the process of tumorigenic transformation of liver progenitor cells (LPCs) (see Fig 1).
We hypothesise that the loss of Arf is a critical early step in the transformation of LPCs. The observation that
Arf is expressed in LPCs is significant as Arf is not expressed in many adult and has also led us to hypothesise
that Arf may be a marker of non-transformed LPCs. We have generated LPCs from embryos of Arf null (-/-)
mice and these cells show an increased propensity to transform (see Fig 2 below). Consistent with this,
karyotype analysis of the Arf-/- LPCs indicates the cells already display signs of
chromosomal abnormalities and instability. Also the culture of Arf-/- LPCs to high density has revealed that the
lack of Arf sensitises the cells to apoptosis and cellular destruction leading us to hypothesise that Arf plays a
significant role in the survival and proliferation of LPCs especially at high density. This project will involve
numerous follow-up experiments aimed at characterising the role of Arf in these processes. This will include
elucidating the role of Arf in regulating the function of key transcription factors such as c-Myc and FoxM1 to
prevent LPC transformation. We are also developing a system for reducing Arf expression utilising lentiviral
shRNAs that will allow us to demonstrate a direct role Arf plays in these cellular processes.
2. The role of the p19Arf tumour suppressor on chromosomal stability in liver-progenitor cells.
With Professor George Yeoh, Biochemistry and Molecular Biology.
We have previously found that the expression of the Arf tumour suppressor is significantly down-regulated to
undetectable levels during the process of tumorigenic transformation of liver progenitor cells (LPCs). We
hypothesise that the loss of Arf is a critical early step in the transformation of LPCs. We have generated LPCs
from embryos of Arf null (-/-) mice and these cells show an increased propensity to transform. Consistent with
this, we have also performed karyotype analysis of the Arf-/- LPCs and the results indicate that the cells already
display signs of chromosomal abnormalities and instability, a key trait of cancer cells. This project will involve
follow-up experiments aimed at determining the role of Arf in preventing chromosomal instability. We
hypothesise that routine culture of LPCs results in chromosomal instability and that loss of Arf and/or culture
under normoxic conditions will hasten this process. Therefore Arf-/- and Arf +/- cells will be serially passaged
under both normoxic and anoxic conditions and cells will be systematically examined with increasing passage
for chromosomal instability. Cells will also be treated with Arf shRNA to reduce Arf levels and together with
parental cells (controls) these will be passaged under normoxic and anoxic conditions
and systematically examined with increasing passage for chromosomal instability. In addition, an inducible
system will be developed to knockdown Arf levels using shRNA. These studies will allow us to determine the
role Arf plays in preventing chromosomal instability and will provide the framework for developing
chromosomal molecular probes for fluorescent in situ hybridisation (FISH) to detect transformed liver stem cells
in mouse and ultimately human liver pathologies.
12
Figure 1: Chromosomal changes seen during transformation of LPC (BMEL) cells at passage 5 (A), 10 (B)
and 15 (C). Large chromosomal abnormalities (upper panels) including loss (red arrowheads) and gain (blue
arrowheads) of genomic material are seen concomitant with transformation of the cells indicated by the ability
of the cells to grow in soft-agar (lower panels). Cells used for growth in soft-agar are the same cells used for
karyotyping at passage 6 (A), 11 (B) and 16 (C). Note the increase in size and number of colonies with
increasing passage number.
Figure 2: p19Arf
expression is lost
during LPC
transformation. A) p19 Arf expression
in non-transformed (NT) and transformed (T) BMEL
and BMOL LPC lines showing Arf expression is lost
during transformation. B) BMEL p19Arf-/- LPCs have
the propensity to form colonies when grown in soft-agar.
Note the increase size and number of colonies compared
with the non-tranformed BMEL A-EGFP (p12) control
cells. Approximately 21% of BMEL Arf null (-/-) LPCs
form colonies in soft-agar at p20.
3. Examining the role of YAP in transformation of liver progenitor cells.
Previous research has established that over-expressing the YAP oncogene in cells results in increased cell
growth and acquired resistance to certain forms of apoptosis, two key traits of cancer cells. We have also
observed that YAP localizes to the nucleolus in non-transformed liver progenitor cells (LPCs) but not in
transformed LPCs. We hypothesise that loss of nucleolus localized YAP results in its activation leading to
cellular transformation. Furthermore, the expression of the p19Arf tumour suppressor is reduced to undetectable
levels in transformed LPCs. Interestingly YAP nucleolus localization is abolished in Arf-/- LPCs suggesting that
Arf is involved in this process. We also hypothesise that Arf and YAP interact biochemically and that
expression of YAP in Arf-/- LPCs will result in an enhanced rate of cellular transformation. This project will (i)
determine whether Arf and YAP interact by performing co-immunopreciptitation (co-IP) experiments; (ii)
examine and compare the effect of YAP over-expression in Arf+/+ and Arf-/- LPCs on the rate of cell growth
and transformation by examining the ability of the cells to grow in low serum and in soft-agar, a key indicator of
transformation and (iii) examine the effect of reactive oxygen species (ROS) on YAP-induced cellular
transformation by culturing LPCs under normoxic and anoxic conditions or by culturing LPCs in the presence of
anti-oxidants, e.g. L-ascorbic acid (vit C) to determine whether ROS contributes to the transforming ability of
YAP.
A
B
C
13
ASSOCIATE PROFESSOR
RETO DORTA Room and Phone available soon (December 2011)
Email: [email protected]
Group webpage: http://www.oci.uzh.ch/group.pages/dorta/home.html
Organometallic Chemistry and Catalysis Our research is directed toward the preparation of reactive transition metal complexes for stoichiometric and
catalytic applications. We focus our attention on the development of new chiral and non-chiral auxiliary ligand
systems which are able to bind, activate and functionalize the substrates at the metal center. The ultimate goal of
the research program is to identify new ligand families and their corresponding metal complexes for new, more
selective or more widely applicable catalytic transformations.
Projects for honours students offer a unique opportunity for getting hands-on experience in modern organic and
inorganic chemistry. State-of-the-art routine lab equipment will be made available and includes synthetic aspects
of the project (Schlenk-line techniques, Glovebox techniques) as well as analytical aspects (GC-MS, GC‟s and
HPLC instruments with chiral stationary phases within the laboratory, NMR and X-ray analysis and other
necessary equipment within the department). The projects will be such as to provide real insights into new
developments in the field of catalyst development and organic synthesis within the timeframe of the honours
degree. Additional related projects will be made available upon request.
PROJECTS
1. Ligand Systems Based on Chiral Sulfoxides and Their Use in Late-Metal Chemistry and Catalysis
several possible projects
Expanding the ligand families capable of acting as
successful entities in metal-mediated reactivity and catalysis
is crucial for future discoveries in this field and will lead to
systems that show unprecedented reactivity patterns. One of
our recent research goals is to identify and apply chiral
chelating sulfoxides as sulfur-based ligands in late-transition
metal chemistry. First results show that these ligands indeed
are able to perform well in a conjugate addition reaction
catalyzed by Rhodium. The honours projects available in
this area of our research will focus on novel ligand systems
of this family and will expand catalytic reactivity to other
reactions catalyzed by late-transition metals. For additional
information on our research, please consult the following
publications: R. Mariz et al., J. Am. Chem Soc. 2008, 130,
2172; J. J. Bürgi et al., Angew. Chem. Int. Ed. 2009, 48,
2768; R. Mariz et al., Chem. Eur. J. 2010, 16, 14335.
R
EWG
chiral Rh/Ir/Pd/Pt cat.
R = H, Alkyl, aryl, OR", NR"2
R' = Alkyl, aryl, OR", NR"2
M = B, Al, Zn, Si, Ti
+ M–NuR
EWGNu
*
R'R'
H+/electrophile
R'
R'
S
S
O
O
R
R
S
S
O
O
R
R
R'
R'
Possible catalytic application:
S
SO
R
O
R''
R,R'' = alkyl, aryl; R' = hydrogen, alkyl, aryl
Fe,Ru
S
R
O
SR''O
Possible disulfoxide ligand structures:
14
2. New Chiral N-Heterocyclic Carbene Ligands in Asymmetric Catalysis
several projects available
Reactions incorporating NHC metal complexes represent some of the most
significant advances in homogeneous catalysis during the last decade, particularly
for alkene metathesis and for coupling reactions. Nevertheless, there is a very
restricted architectural choice for these ligand system and this is particularly
hindering development of chiral monodentate NHCs. In the last few years, we
have therefore initiated a research program that proposes the synthesis of new
classes of monodentate, chiral NHCs that incorporate substituted naphthyl
sidechains on the nitrogen atoms. In doing so, we are indirectly relying on a very
successful design motif in chiral ligand synthesis that goes back to Noyori‟s bis-
phosphine ligand BINAP. These new types of ligand systems will allow for the
synthesis of new transition metal complexes, where our focus will particularly lie
on the isolation of highly unsaturated precatalysts. Special emphasis in subsequent
applications will be put on the identification of more active chiral rhodium and
iridium NHC compounds in catalysis, development of better asymmetric nickel,
palladium and ruthenium mediated transformations and the development of
unknown NHC-Ag catalysis. For preliminary data from our group on this project,
see: X. Luan et al., Org. Lett. 2008, 10, 5569; X. Luan et al., Org. Lett. 2010, 12,
1912.
3. New Catalysts and New Substrates in Ruthenium-catalyzed Metathesis Reactions
Olefin metathesis has experienced a significant evolution in the last
decades and is becoming one of the most useful synthetic transformations
for generating carbon-carbon double bonds. The reaction can be applied in
a great variety of synthetically useful permutations that include ring-
closing metathesis (RCM), cross metathesis (CM) and enyne metathesis.
Among the catalysts that have been developed, ruthenium alkylidene
complexes incorporating an N-heterocyclic carbene (NHC) ancillary ligand
(Grubbs‟ second-generation catalyst) have become the most widely used in
organic synthesis. The goal of this project is twofold; we have already been
able to show that modifying the NHC ligand (see project 2) can bring about
a clear increase in catalyst performance; further fine-tuning is therefore a
worthwhile target and is also expected to lead to new reactivity in difficult
or novel applications of the metathesis reaction. Indeed, metathetical
reactivity of relatively electron-rich double bonds is still very challenging,
presumably due to the fact that stable, Fischer-carbene type complexes are
generated upon reaction with the catalyst‟s metal center. Here, new results
in our group have shown that modification of the substrates themselves
might lead the way to exploiting metathesis reactions that have previously
not been known. For recent results from our group, see: X. Luan et al., J.
Am. Chem. Soc. 2008, 130, 6848; M. Gatti et al., J. Am. Chem. Soc. 2009,
131, 9498; M. Gatti et al., J. Am. Chem. Soc. 2010, 132, 15179.
N N
C2-symmetry
R1
R2
R2
R1
N N
R1
R1
R1R2R2
R1
Successful NHC ligands
Proposed Chiral NHC ligands
X = Halide, OR, NR2 etc.
RuCl
Cl
OiPr
Ph
N N
R7
R2
R2
R7
Y Y
XX
n nRu cat.
CR2
Ru cat.
+
X
CR2
X
R+
R+
Ring-closing metathesis
Cross metathesis
Catalyst:
15
DR ELA EROGLU RESEARCH ASSISTANT PROFESSOR
Centre for Strategic Nano-Fabrication and
ARC Centre of Excellence in Plant Energy Biology
Room 3.41, Bayliss Building, Phone: 6488 2558
Email: [email protected]
My research projects involve photosynthetic microorganisms (such as microalgae and photosynthetic bacteria)
and their applications to nanotechnology. The following topics cover several ―green‖ bioprocesses while
combining several interdisciplinary fields including Biotechnology, Nanotechnology, Agricultural and
Environmental Sciences
PROJECTS
1. Wastewater treatment processes with Immobilized Algae
with Prof. Steve Smith (CoE in Plant Energy Biology), Prof. Colin L. Raston, and Dr. Swaminathan Iyer
Wastewater treatment is the process of eliminating unwanted chemicals, or biological contaminants from the
impure water. It mainly includes liquid wastes released by houses, industrial properties, and/or agricultural
processes; while having a wide range of contaminants at various concentrations (Metcalf and Eddy 2003). As a
relatively recent bioprocess, microalgal cultivation in wastewaters has a combination of several advantages such
as integrated wastewater treatment and simultaneous algal biomass production, which can be further exploited
for biofuel production (in the form of biodiesel, biohydrogen, or biogas), food additives, fertilizers and soil
conditioners, cosmetics, pharmaceuticals, and many other valuable chemicals (Mallick 2002). Microalgae are
the recent organism of choice for the renewable generation of hydrocarbon-based biofuels, with high biofuel
yields in comparison with plant-oils (Eroglu and Melis 2009). Microalgae have several other advantages as it
can grow within short time intervals, does not require many resources to produce, and can be utilized for the
reduction of CO2 emissions by using carbon dioxide for biomass and/or energy production. In addition to the
utilization of the wastewater contents for algal biomass formation, the dissolved oxygen released by the algae is
also useful to oxidize waste organic matter.
One of the main problems to obtain a productive algal water
treatment and bioenergy systems is the harvesting, dewatering
and processing of algal biomass. In this project novel algal
immobilization approaches will be adopted by employing
various nanotechnological strategies, such as electrospinning.
This work has great prospects to combine interdisciplinary fields
in the national and international collaborations. Several
industrial waste treatment plants, and the Governmental or
private water-cooperation are potential end-users of the
developed-technology. Image: http://www.westernenvirosolutions.net/
16
2. Nutrient Recovery for the Generation of Sustainable Biofertilizers
with Dr.Sasha Jenkins (from the School of Earth and Environment)
Several effluent wastewaters (i.e., agricultural, municipal, industrial) contain high amounts of nitrate and
phosphates that needs to be removed from their effluents before discharging into their environment. The
exposure of surrounding system and groundwater to pollution brings severe environmental regulations to be
imposed on these industries. Removal of these nutrients is very essential especially to avoid eutrophication of
the surrounding water sources that can result several environmental impacts. Nitrate and phosphate uptake can
be achieved via algal pond systems, while the algal biomass is
harvested and can be recycled as a fertilizer.
In this study, we‟ll be investigating the treatment of effluent wastes
with high nitrate and phosphate loadings by immobilized algal
cultures. Then the algal cultures will be harvested and the immobilized
algal biomass will be recycled on land as “slow-release” fertilizer
which will be highly beneficial for organic farming. Goal of this
research is to develop and operate a sustainable nutrient recovery
system from various wastewaters by using immobilized algal systems
and mixing these algae-hydrogel combinations with the soil as a
moisture-rich fertilizer and soil enhancer. Nitrate and phosphate
recovered from the effluent wastewater will be recycled back to soil
by algae, whereas hydrogel matrix can also be beneficial for providing moisture to the soil. Image: http://www.biosynherb.com/bio-fertilizer.html
3. Biosynthesis of nanoparticles
with Prof. Steve Smith (CoE in Plant Energy Biology), Prof. Colin L. Raston, Dr. Swaminathan Iyer, and Dr.
Jeremy Shaw (from Centre for Microscopy, Characterisation and Analysis)
Metal nanoparticles have recently been receiving significant interest, due to their distinctive chemical, magnetic,
electronic, and optical properties. As a result of their high surface-to volume ratio, they have been used for
various applications such as catalysis, biological labelling, electronics, and optical devices (Lengke et al. 2007).
Rather than following the conventional chemical pathways, biological materials can be used for the synthesis of
metal nano-particles as ecological stabilisers.
For this context, several microorganisms will be investigated for their nanoparticle (such as Palladium
nanoparticles) production capability. Transmission Electron Microscopy (TEM) techniques will be developed
and applied for the imaging and the characterization of nanoparticles.
Image: Gillian Walters and Ivan P. Parkin (2009) J. Mater. Chem., 19, 574
References
Lengke et al. (2007). Langmuir, 23, 8982-8987
Eroglu and Melis (2009) Biotechnology and Bioengineering, 102(5): 1406-1415
Mallick (2002) Biometals, 15: 377–90
Metcalf and Eddy, Inc (2003) Wastewater engineering: treatment and reuse. 4th
ed., McGraw-Hill, New
York.
17
DR GAVIN R FLEMATTI ARC Postdoctoral Fellow
Room 4.17, Bayliss Building, Phone: 6488 4461
E-mail: [email protected]
Research Interests
My main research interest is in the field of bioactive natural products. I work closely with A/Prof Emilio
Ghisalberti in this regard and together with collaborators from other disciplines we are interested in the
detection, isolation and identification of natural products that demonstrate some form of biological activity.
Some possible honours projects are summarised below which I am happy to discuss further.
PROJECTS
1. Isolation of bioactive compounds that reduce methane emission in ruminants.
Collaboration with A/Prof Phil Vercoe, School of Animal Biology, UWA
In Australia, 90% of the total greenhouse gas emissions from agriculture stem from gases produced as a natural
end-product of the digestion in ruminants (sheep and cattle), including methane as the most potent greenhouse
gas. One way to reduce methane emissions from animals is to feed them plants that contain naturally occurring
secondary compounds with antimicrobial properties that can inhibit methanogenic microorganisms in the rumen.
A/Prof Phil Vercoe‟s research group at UWA Animal Biology has identified several Australian native plants
with these antimethanogenic properties in the rumen. However, the chemistry, the metabolism in the rumen and
the mode of action of these compounds is unclear.
This project aims to isolate and identify the major secondary metabolites from selected Australian native plants
and investigate their role in reducing methane production by the rumen microbes.
2. Investigation of volatile organic compounds emitted from Australian truffles.
Collaboration with Prof Garry Lee, Centre for Forensic Science, UWA
Truffles are subterranean edible fungi that traditionally grow in various parts of Europe, particularly in Italy and
France. They are highly appreciated due to their characteristic aroma and are used mainly uncooked in French
and Italian cuisine, particularly the black perigord truffle (Tuber melanosporum). Previous research has
identified over 200 volatile compounds that are emitted from truffles, including many alcohols, ketones,
aldehydes, aromatics and sulphur compounds.1
Studies show that the geographical location plays a significant role on the composition of the truffle volatiles.2
To date, there have been no reports on the composition of volatiles from Australian grown truffles. This project
will investigate the volatiles emitted by black truffles (T. melanosporum) at various stages of maturity from at
least three different locations in Australia (Western Australia, New South Whales and Tasmania). Commercially
available truffle oils will also be analysed and compared with fresh samples using solid phase micro-extraction
(SPME) and GC/MS. The purpose of this work will be to identify volatiles that allow differentiation of truffles
grown from different regions and at different stages of maturity.
Refs: 1 Cullere, L., Food Chemistry, 122, 300-306 (2010).
2 Gioacchini, M. A., Rapid Communications in Mass Spectrometry, 22, 3147-3153, (2008).
18
DR SIMON GRABOWSKY Room 4.29, Bayliss building, Phone: 6488 3515,
Email: [email protected]
The aim of our research is to find the exact locations of electrons within molecules and make them visible. We
can use the electron density, which tells us about electron concentration and depletion, and we can use electron
localisation functions, which tell us where electron pairs are localised. If we know the exact distribution of
electrons within molecules and where they preferably pair up, we can derive information about chemical
bonding and reactivity.
We use two ways to extract the desired information: an experimental one and a theoretical one. X-ray diffraction
experiments on single crystals at very high, i.e. sub-atomic, resolution and at ultra-low temperatures (down to
8K) allow us to obtain the electron-density distribution of the scrutinised compound. New techniques go even
beyond this and allow to derive an experimental wavefunction from the X-ray diffraction data, which can be
used to calculate electron localisation functions additionally to the electron density. The theoretical way uses
quantum-mechanical ab-initio calculations on the computer to derive all necessary information from a theoretical
wavefunction. Applications of these techniques are widespread: On the one hand, we are interested in
compounds suitable for drug design and study potential active centres for interactions with enzymes; on the other
hand, we are interested to shed light on unusual bonding situations in organic and inorganic compounds.
PROJECTS
1. Comparing the electronic nature of different potential protease inhibitors to facilitate drug design
Proteases are enzymes that catalyse the hydrolysis of
peptide bonds. They are essential for any organism
in many different ways. But they also play an
important role in the dissemination of cancer.
Tumour cells release pathogenic forms of
autologous proteases like cathepsin or collagenases.
Therefore, drugmakers search for compounds which
can irreversibly inhibit specific proteases.
Our collaboration partners (University Wuerzburg,
Germany) synthesise compounds which attack
cysteine or asparagine groups in proteases via an
electrophilic attack at the mercapto (SH) group in
the case of cysteine or at the carboxyl group
(COOH) in the case of asparagine. We have already
measured high-resolution X-ray diffraction data sets
of three of these compounds with different active
electrophilic centres (an epoxide group, an electron-
deficient double bond, a sulphur-containing five-
membered ring). See the figure for the electrostatics around an epoxide model compound.
In this honours project, we aim to measure a fourth high-resolution data set at the in-house X-ray
diffractometer and evaluate the crystal electron density as well as electron-pair localizability within these
four protease-inhibitor model compounds. A detailed comparison of the different active centres at the
electronic level will give insights into the inhibition mechanism and thus the usefulness for drug design.
19
2. Transferability of sub-molecular properties in the electron density
The electron-density distribution for any molecule
can be subdivided into functional group or even
atomic regions. This is part of Richard Bader‘s
Quantum Theory of Atoms in Molecules. Physics
predicts that these sub-molecular fragments are
transferable between different molecules. The
figure shows a cut-plane through the electron
density of two fused rings. Regions in the electron
density belonging to the individual atoms can be
identified. You can imagine how these regions
could be extracted from the picture using a scalpel
and could be glued together in a different
arrangement to construct the electron density of a
different compound. In fact, the total electron
density of a compound, e.g. a large one like a
protein which cannot be measured to high
resolution, can be built up from atomic fragments
like a three-dimensional puzzle.
This concept is extremely useful and has led to the
development of electron-density data banks which
store atomic electron-density building blocks. However, this has only been tested for and has been applied
within the so-called multipole expansion of electron-density modelling. But we want to test this in a more
general way and for more functions than only the electron density.
We have measured the high-resolution X-ray diffraction data sets of six different tripeptides of the type L-
alanyl-X-L-alanine in the past where X is a variable amino acid. The aim of this honours project is to use these
data sets to extract experimental wavefunctions and to compare derived properties with respect to transferability
of sub-molecular fragments.
Other projects may be available after consultation
For an introduction to these research areas, see the following publications:
P. Luger, Fast electron density methods in the life sciences – a routine application in the future? Org.
Biomol. Chem. 2007, 5, 2529-2540.
T. Koritsanszky, P. Coppens, Chemical Applications of X-ray Charge Density Analysis. Chem Rev.
2001, 101, 1583-1627
Watch the lecture on his website http://www.chemistry.mcmaster.ca/bader/
A. Savin, R. Nesper, S. Wengert, T. F. Faessler, ELF: The Electron Localization Function. Angew.
Chem. Int. Ed. Engl. 1997, 36, 1808-1832.
S. Grabowsky, T. Pfeuffer, W. Morgenroth, C. Paulmann, T. Schirmeister, P. Luger, A comparative
study on the experimentally derived electron densities of three protease inhibitor model compounds.
Org. Biomol. Chem. 2008, 6, 2295-2307.
S. Grabowsky, T. Schirmeister, C. Paulmann, T. Pfeuffer, P. Luger, Effect of Electron-Withdrawing
Substituents on the Epoxide Ring: An Experimental and Theoretical Electron Density Analysis of a
Series of Epoxide Derivatives. J. Org. Chem. 2011, 76, 1305-1318.
S. Grabowsky, R. Kalinowski, M. Weber, D. Foerster, C. Paulmann, P. Luger, Transferability and
reproducibility in electron-density studies – bond-topological and atomic properties of tripeptides of
the type L-alanyl-X-L-alanine. Acta Cryst. B 2009, 65, 488-501.
20
WINTHROP PROFESSOR
PETER HARTMANN Room 2.03, MCS Building, Phone 6488 3327
Email: [email protected]
Human Lactation
Winthrop Professor Peter Hartmann leads a large research group that carries out both basic and applied lactation
research with women and infants. Despite a plethora of evidence showing breast milk is the best nutrition many
women fail to sustain exclusive breastfeeding for 6 months as recommended by WHO. The aim of this group is
to provide an evidence base for clinical protocols and management of lactation difficulties. To achieve this
objective a fundamental research into the physiology and biochemistry of milk synthesis milk secretion, cell in
milk (immune and stem), milk ejection, the mechanics of breastfeeding and the control of infant appetite is
carried out.
The following projects will increase the knowledge base of lactation substantially and are available to honors
students
PROJECTS 1. Reactive oxygen species in human milk with Dr James Lui and RA/Prof Ching Tat Lai
Reactive oxygen species (ROS) have received much attention due to their high reactivity and ability to modify
other biomolecules. These modifications may potentially be so devastating that they precipitate damage to tissue
and subsequently cause disease. ROS can be generated at the cellular level as well as during environmental
stress (e.g. ultraviolet irradiation, ultrasound or heat exposure). Although the human lactating breast produces
high quantities of antioxidant proteins and molecules that scavenge these ROS, recent evidence suggests that
ROS may function as antimicrobial agents. We have tested several assays to detect ROS in human milk and
preliminary results have identified ROS. This project will extend this work with the aim of extensively
documenting ROS and determining their role in human milk.
2. Peptide profile of human milk with Dr James Lui
Certain classes of peptides in human milk have been
shown to have bioactive functions such as providing
infant immunity and stimulating infant growth. In
addition antimicrobial properties have been
demonstrated in in vitro experiments. Data from
previous studies have only considered either one or
several specific groups of peptides derived from
protease-digested proteins found in human milk. It is
well known that large variations in the components of
human milk exist between lactating mothers therefore it
is impossible to draw firm conclusions about the
bioactive functions of these peptides. This project will
endeavor to characterize the natural peptide profile of
human milk at different stages of lactation thus providing a fundamental understanding of the involvement and
significance of peptides in human milk with regard to both the mother and the infant.
21
Mass spectrometry data and PCA analysis
3. Bacteriostatic properties of human milk with RA/Prof Ching Tat Lai
While breastfeeding is recommended first and foremost by WHO there are many situations where the mother
needs to express her milk to be fed to the infant for instance premature infants are too ill and weak to breastfeed.
Since breastmilk improves preterm infants short and long term health outcomes it is imperative that they receive
this milk safely. Human milk is unique in that it possesses bacteriostatic properties that are apparent when milk
is stored over time. These properties are diminished when the milk is pasteurized prior to feeding. Despite the
importance of this property to the infant little investigation has been carried out in this area. This project is
designed to further investigate the bacteriostatic effects of milk and to determine which components are
responsible for these effects.
4. Cellular biochemistry of human milk with Dr James Lui and RA/Prof Ching Tat Lai
Recent research indicates the existence of cell
population, intact cellular organelles and bacteria in
expressed breast milk. Although current studies are
beginning to characterize the different cell types and
bacteria existing in breast milk, our understanding of
the functional significance of the cells, organelles and
bacterial populations in breast milk are still unclear.
Biochemical contribution of these populations to the
milk could be a window of opportunity to observe
physiological changes in the lactating breast. This
study will take a focus approach to explore cellular
biomolecules in breast milk to determine the
functional relationship between these populations in
breast milk. This may lead to a way of monitoring
any changes in the health of the mother, which may
inadvertently affect the health of the newborn infant.
These projects provide exciting insights into the components of breast milk and their functions. They
provide important knowledge that will contribute to the development and refining of optimal storage
conditions for the milk. Ultimately it may be possible to tailor components in the milk to benefit ill and
premature infants.
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ASSOCIATE PROFESSOR
DYLAN JAYATILAKA Room 4.30, Bayliss building, Phone: 6488 3138,
Email: [email protected]
Theoretical and Computational Chemistry
I am interested in a number of areas, including:
Quantum chemistry: using quantum mechanics to calculate molecular properties e.g. shapes, dipole
moments, polarisabilities. We use existing computer programs and we write our own too.
Chemical concepts from quantum mechanics. Although quantum mechanics can produce properties, by
following the rules, it is often difficult to understand and interpret these properties in terms of “atoms”
and “bonds” and all the usual terms that chemists use. I'm interested in developing theories and
methods to do this.
Crystallography and diffraction: I'm interested in using diffraction experiments to improve quantum
chemistry calculations, and vice versa, using quantum chemistry to improve measurements from X-ray
and polarized neutron diffraction experiments.
Development of reusable software. I have written a program library called Tonto which makes
developing new quantum chemistry and crystallography methods easier than normal.
Visualisation of complex chemical data. I have helped developed a program called Crystal Explorer to
visualize crystal structure packing information in high quality 3D graphics.
What do I need to know to do a computational project?
You need to be familiar with computers (who isn't) and if not, you need to be good at maths. You will develop
skills in dealing with Unix computers in the projects to run existing programs. For more specialist projects, you
will need to be interested in writing programs to solve problems. A general facility with numbers helps.
PROJECTS
1. Why do crystals have the shape they do? What about the spaces between molecules in crystals?
With: Prof. Mark Spackman, Dr Mike Turner
The physical properties of a crystal are directly connected to the way the
molecules pack to form a crystal. Why do molecules pack in one way, and not
another? Even though we know the underlying laws, this is a basic question
that is still largely unanswered. A unique “fingerprint” can be made of this
surface (shown left, for urea). The fingerprint is
easier to see and understand since it is two
dimensional. In this project, we want you to
systematically examine the fingerprints for
many structures and to try and compare them to
see if we can at least classify all the different
types of crystal structure. You could also
develop a new kind of fingerprint which
characterises the void spaces between molecules. The voids between molecules
are useful in themselves, because other molecules (such as hydrogen) can be
fitted into them for storage.
Further reading: Spackman MA, Jayatilaka D (2009) Hirshfeld surface analysis. Cryst. Eng. Comm. 11:19-32.
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2. What defines a bond? Bond indices from quantum mechanics.
Have you ever wondered when you are allowed to draw two lines between an atom in a molecule and call it a
bond? Recently we published a paper that allows to calculate the bond index between two atoms in a molecule
from the wavefunction of the molecule. Furthermore, we showed how to calculate an ionic bond index and a
covalent bond index. We've tested the method on a few systems, and the results are mostly very good, but there
are a few anomalies, especially for group II elements. In this project you will use our programs and calculate
bond indices for a range of interesting systems. We are particularly interested in performing calculations on a
recently reported compounds which is claimed to have a quintuple bond. You will try to discover the reason for
the anomalous results.
Further reading: Gould MD, Taylor C, Wolff SK, Chandler GS (2008) A definition for the covalent and ionic
bond index in a molecule An approach based on Roby's atomic projection operators. Theor. Chem. Acc.:275-
290.
3. Visualising energy densities in molecules
The energy of a molecule is a crucial property. However, the energy is a
property of the whole molecule: so where exactly is the energy located in
the molecule? In this project we are interested in obtaining properties such
as the energy density in the molecule. Prof. Gibbs and I have found new
expressions for a number of property densities (kinetic energy density,
potential energy density, and so on) obtained from wavefunctions. None of
these properties has ever been plotted before. In this project you will
investigate these new energy densities (there are 48) for a range of simple
molecules. Shown to the right is the plot of the electron localisation function
for urea (this is not one of the ones you will look at). This will be a
colourful project.
Further reading: Grimwood DJ, Bytheway I, Jayatilaka D (2003) Wave functions derived from experiment. V.
Investigation of electron densities, electrostatic potentials, and electron localization functions for
noncentrosymmetric crystals. J. Comp. Chem. 24:470-83.
4. 3D and dome visualisation with Crystal Explorer
With: Prof. Paul Bourke, Dr. Mike Turner
The ability to visualise and interpret crystal structures is an important
aspect of structural science. We have developed a program called Crystal
Explorer used to visualise crystal structures, Hirshfeld surfaces, and
Hirshfeld surface fingerpints (see project 1). Crystal Explorer is popular
and widely used. In this project you will extend Crystal Explorer to
display on a dome projector to aid visualisation (shown left). You will
also try to convert the program to display in 3D, using 3D glasses. This
project will require some background in programming or a strong desire
to develop skills in this area. It will be co-supervised with Paul Bourke
from the WA supercomputing facility (WASP), which has dome and 3D
projection facilities.
24
PROFESSOR
GEORGE KOUTSANTONIS Room 3.11, Bayliss building, Phone: 6488 3177,
Email: [email protected]
Metals in Chemistry and Nanochemistry Our group is interested in the role of metals in functional materials. While the role played by metals in materials
is still evolving and there is a an increasing effort to incorporate redox–active centres into many materials, e.g.
conducting polymers, in an effort to create highly efficient redox conductivity for sensor, catalytic,
photochemical and photoelectronic applications. We are participating members of the WA Centre of Excellence
in Nanochemistry.
PROJECTS
1. Biomimetic Complexes of the Mg/Ca oxide cluster of Photosystem II
The inorganic cubane complex known as the manganese-calcium oxide cluster,
commonly referred to as the "Oxygen Evolving Complex" or OEC (also referred to as
a photosynthetic water oxidase). The OEC is located on the oxidizing side of
Photosystem II (PSII), and isolated within chloroplasts, a plastid found in all plants
and algae. The OEC is also found in one group of bacteria, the Cyanobacteria. It is
believed that the Cyanobacteria are the endosymbiotic ancestors of modern day
chloroplasts. At the active site water oxidation procceds at a pentanuclear Mn4Ca
oxide particle with an “organic sheath” protecting the core. An attractive method for
the formation of nanoparticles derived from metal compounds is the use of a
particular ligand to excise clusters from the lattice of a simple species. More commonly, however, the "excision"
is a formal process, in that while the cluster may be recognisable as a portion of an extended lattice, it is not, in
fact, formed by direct fragmentation of that lattice..
In this project we will utilise polyphenolic compounds, called calixarenes, as a template to build Mn/Ca
clusters upon and to introduce the geometric constraints required for the enzyme function to be mimicked.
2. Molecular Computing: Dihydropyrene-based organometallic molecular switch
with Assoc. Prof. Matthew Piggott)
Dimethyldihydropyrenes are fascinating molecules with a planar 14- -electron periphery, making them
aromatic. They are easily converted to their valence tautomers, cyclophanedienes, by irradiation with certain
wavelengths of light. This project will involve the synthesis of a novel diethynyl-substituted dihydropyrene that
will be used to prepare organometallic complexes. Switching between the valence tautomers is expected to
drastically change the conductivity of the organic ligand, which in turn will affect properties such as colour,
crystallinity and redox potential.
We will prepare metal complexes of dimethyldihydropyrenes that will have the potential to fine tune the
physical properties of these materials for application in new computing technologies.
3. Redox-active Metallomicelles
Metallosurfactants are an emerging class of materials which offer interesting alternatives to traditional “organic”
surfactants due to the range of properties
inherent to complexed metal ions
Introduction of such a centre can impart the
magnetic and electronic properties, as well as
the redox and catalytic activity of the
complex to the surfactant system, which of
course can be concentrated at an interface, be it polar/apolar (e.g. micelles,
vesicles), solid/liquid (e.g. monolayers) or liquid/gas (e.g. Langmuir-Blodgett
films). Cationic surfactants have general applications such as biocidal agents, and
there has been recent interest in their use as DNA delivery agents for gene therapy.
We have shown that copper and cobalt metallosurfactants can form wormlike
25
micelles in aqueous solution which may co-exist with, or easily interconvert with vesicle structures. The
cylindrical micelle structures are of nanometer dimensions and these cylindrical structures are unusual for triple
chain surfactants, not easily accounted for using geometrical packing arguments. The solution behaviour has
been characterised by cryo-TEM and SAXS measurements. Both the Cu and Co compounds display viscoelastic
solutions at 10 wt% which coupled with the wide variety of stable metal complexes formed by the cage head
group augur exciting materials for possible application in the production of mesoporous silica structures loaded
with metal aggregates for a variety of catalytic applications.
4. Redox-active Metal Complex Oligiothiophenes as Sensors and Devices
with Dr Gavin Collis, CSIRO Material Science and Engineering
The drive for new devices that have utility in electrochemical sensing
applications or for clean electrocatalysis has seen considerable effort
expended in the modification of electrodes. Two intensely studied
approaches to construct of such electrodes has been the formation of
self-assembled monolayers or by the deposition of a funtionalised
polymer on the electrode surface. In this latter case the most widely
studied class of monomers for the production of polymer modified
electrodes are functionalised thiophenes and oligothiophenes, as they
can be readily electropolymerized directly onto the electrode.
We have
recently shown
that
oligiothiophenes that have metal complexes directly
attached to the polymerisable unit have difficulty in
beingpolymerised. Thus this project will strive to prepare
new monomers for polymerisation that have variable
linkers for attachment to metal complexes. The targets in the first instance are shown adjacent.
5. Charge density analysis of fundamental host-guest supramolecular systems
several projects, with Prof Mark Spackman and Dr Alex Sobolev, UWA
Although supramolecular chemistry is one of the most active fields of modern chemistry, very
little seems to be known about the detailed nature of the host and guest
systems that comprise these aggregates. Supramolecular systems – molecular
aggregates – underpin the design and development of materials in areas as
diverse as catalysis, targeted drug delivery, gas storage, chemical separation
and nonlinear optics. They also serve as models for complex phenomena such as self-assembly
and ligand-receptor binding. Projects in this area are part of a research program aimed at a greater understanding
of intermolecular interactions and the properties of host-guest systems in the solid state, particularly organic
clathrates and complexes formed by small molecules interacting with crown ethers, calixarenes, molecular
tweezers and cages (some examples are given in the figure below). These projects
will involve some synthesis, and measurement of highly accurate X-ray diffraction
data, complementary neutron diffraction experiments, quantum chemical
calculations and computer graphics. A particular focus of the charge density
analyses will be the polarization and dipole moment of guest molecules as a
function of the changing electrostatic nature of the host systems.
6. New Organometallic Materials with Assoc Prof Matthew Piggott)
Photocatalysis and its application to solar energy conversion is an important research problem for the next
century particularly in light of the peak oil problem that faces
current energy generation strategies.
This project seeks to prepare new metallotectons with the ability to
potentially control energy and electron transfer processes. One
way in which to do this is to recruit pendant or bridging aromatic
groups for this purpose and a readily available moiety for this is the
pentacene unit. Aromatic units of differing structure will allow us
to control the HOMO-LUMO and band gap. There is a significant
synthetic component involved in this project the majority of which
is supported by solid literature procedures.
The molecule in blue will allow us to target additional allenylidene complexes with interesting
properties and the molecule in red will allow a systematic investigation on metal-ligand combinations and their
effect on the electronic properties of the complexes.
OHC
OHC
O
O
+KOH
O
O
Pri3SiCCLi
HO
HO
C2SiPri3
C2SiPri3
SnCl2
C
C
C
C
SiPri3
SiPri3
C CC C[M] [M] C CC C[M] [M]
S S
S
HN
M
S S
S
HN
MO
M = metal complex
26
ASSOCIATE PROFESSOR
MARTHA LUDWIG On study leave July-December 2012
Room 3.05, Bayliss Building, Phone: 6488 3744
Email: [email protected]
The Molecular Evolution of Photosynthetic Pathways
Terrestrial plants are typically grouped according to the biochemical pathway they use to fix atmospheric CO2
into carbohydrates – the so-called C3 plants, which include crop species such as rice and wheat as well as nearly
all trees; the C4 plants, which include crop plants like corn and sugarcane, and some of the world‟s worst weeds;
and the Crassulacean Acid Metabolism (CAM) plants, which include cactuses, orchids and pineapple. C4 and
CAM plants evolved from C3 plants, and some groups of plants have left “evolutionary footprints” that give us
insights into how this process has occurred at the molecular level. Many CAM plants are able to “switch”
between pathways, depending on the environmental conditions and/or their developmental stage.
Harnessing the photosynthetic biochemistry of C4 plants for increased food, fodder and fuel –
supercharging C3 plants
The global demand for cereals, which are major food sources for animals including humans and are important in
the biofuels industry, has been forecasted to increase by 60% for 2050, and with consumption being greater than
production in seven of the last nine years, and 2008 stockpiles at 70 days of global consumption, major
challenges face agricultural sectors and governments with respect to food, feed and fuel securities. Increasing
productivity is unlikely to be accomplished only by conventional breeding methods. A second “green
revolution” that includes biotechnology is inevitable for some crops and regions. The higher photosynthetic
rates, greater efficiency in the use of water and nitrogen of C4 plants relative to C3 plants in arid and saline
environments – environments that are expanding in many parts of the world due to global climate change – are
desirable traits, which if introduced into C3 plants, have the potential to increase yield. In other words, we are
looking to “supercharge” C3 crops like rice and wheat by giving them a C4 pathway.
Toward this objective, a major aim of the work in the lab is to understand the molecular biology, biochemistry
and cell biology of the enzymes in the C4 photosynthetic pathway. This includes the identification of the control
regions of the genes coding for these enzymes. Such information will be used to make informed and strategic
decisions regarding the transfer of particular C4 enzymes, or an entire C4 pathway, into C3 plants to increase
yield while restricting negative impacts on the environment.
We are using tools of cell and molecular biology and molecular genetics such as differential cDNA library
construction and screening, quantitative reverse transcription PCR (qRT-PCR), transcriptome sequencing, and
immunocytochemistry to identify key proteins involved in the above processes and examine the expression
patterns of their genes. These studies will give insight into the evolution of photosynthesis, the process on which
all life depends, and the plasticity of plants in obtaining nutrients and water from their environment. This
information will open avenues for manipulating these pathways in economically valuable plants and will
increase our knowledge of how plants may respond and cope with predicted future climate scenarios.
PROJECTS
The plants we use in our work are in two evolutionarily significant genera – Flaveria and Neurachne, the latter
being native to Western Australia, and only found in Australia! These groups of plants are important model
systems for examining molecular evolutionary questions because the individual species in the genera use the C3,
C4 or an intermediate C3-C4 photosynthetic pathway and represent a living continuum from the ancestral C3
condition to the evolutionarily advanced C4 state. This allows us to discover the changes that occurred during
the evolution of the C4 pathway at the level of the genes, transcripts and the proteins they encode. This involves:
1. Comparison of gene expression patterns of key enzymes in C3, C4 and intermediate C3-C4 species of
Flaveria and/or Neurachne using qRT-PCR, transcriptome sequencing and/or in situ labeling techniques.
2. The biochemical characterisation of photosynthetic isoenzymes that function in the same intracellular
compartment, and the identification of the proteins with which they interact.
27
3. Identification of regulatory regions that control the expression of genes encoding photosynthetic enzymes.
4. Exploring potential correlations between ploidy and survival under biotic and abiotic stress conditions.
28
ASSOCIATE PROFESSOR
THOMAS MARTIN Room 3.47, Bayliss Building, Phone: 6488 3331
Email: [email protected] UH
The Signalling and Protein Interaction Group
We are interested in cellular signalling and how this impacts on plant development and function. Learning about
this will help us to identify mechanisms by which plants can be improved to be for example drought, salt or
stress resistant or to generate higher yields. These are desirable traits for plants growing under the harsh
environmental conditions in Australia.
To this end we investigate two gene families related to stress responses in plants:
a) One is a class of histone deacetylases (HD2) found only in plants. These are proteins involved in the
regulation of gene expression by deacetylation of histones which causes changes in chromatin structure.
Some of these plant specific histone deacetylases were reported to lead to increased drought and salt
tolerance when overexpressed in Arabidopsis (1).
b) The other is a family of nitrilases which are potentially involved in cyanide detoxification and plant
hormone biosynthesis (2).
Using a state of the art protein interaction system named Bimolecular Fluorescent Complementation (Fig 1) we
have shown that members of the plant specific histone deacetylases and the nitrilases interact with 14-3-3
proteins (Fig 2 a and b). These 14-3-3 proteins bind to other proteins and regulate their activity, cellular
localisation or stability in response to intracellular or extracellular signals and thereby impact on protein
activities and functions (3). Our interest is to understand what the impact of this regulatory interaction between
14-3-3 proteins and histone deacetylases and 14-3-3 proteins and nitrilases is and how this contributes to normal
plant function, especially under stress conditions.
Figure 1: The principle of Biomolecular
Fluorescence Complementation (BiFC). Two
non-fluorescent parts of the Yellow Fluorescent
Protein (YFP) are fused to two proteins assumed
to interact, for example a 14-3-3 protein and a
potentially 14-3-3 regulated protein (A and B). If
these proteins do not interact (left) we will not
observe fluorescence. Interaction of A with B
(right) reconstitutes a functional YFP and
fluorescence can be observed using fluorescence
microscopy The great advantage of this system is
that it can be used in living plants instead of
looking at interactions in vitro. (from 4).
Figure 2: Interaction of 14-3-3 proteisn with histone
deacetylase (a) and nitrilase 1 (b) demonstrated using
Biomolecular Fluorescence Complementation (BiFC). (a) 14-3-3 mu was tested for interaction with the plant
specific histone deacetylase HD2C. Interaction was
found to occur in the nucleus (N) and nucleolus (No).
(b) Interaction of 14-3-3 proteins with nitrilase 1 after
induction of cell death. The interaction occurs usually
in the cytoplasm of plant cells but localises to round
structures after cell death induction as shown in figure
b.
a b
29
PROJECTS
1. Investigating the regulation of plant specific histone deacetylases by 14-3-3 proteins
Histone deacetylases regulate gene expression by deacetylating histones thereby leading to changes in chromatin
structure. We are interested in a subfamily of histone deacetylases found only in plants some of which were
reported to lead to increased drought and salt tolerance when overexpressed in Arabidopsis (1). The degree of
salt and drought tolerance caused by overexpression can potentially be increased significantly by preventing the
regulation of histone deacetylase activity caused by interaction with regulatory proteins such as 14-3-3 proteins
which we identified (Figure 2a). Removing such regulation would potentially allow generating plants which are
better able to cope with stresses such as salt and drought stress. We postulate that preventing 14-3-3 binding to
histone deacetylases will increase the enzymes activity or prevent its inactivation. Overexpressing such mutated
histone deacetylases, i.e. those which are not controlled on the protein level, may in turn increase tolerance of
plants to stress conditions such as high salt and drought. The honours project will explore the regulation of
histone deacetylases by 14-3-3 proteins.
The aims of this project are:
a) To identify and mutate 14-3-3 binding sites in histone deacetylase HD2a and HD2b
b) To verify the loss of protein interaction in living plant cells using Bimolecular Fluorescence
Complementation
c) To test the mutated enzymes for changes in enzymatic properties and regulation.
2. Identifying novel protein interactions of plant specific histone deacetylases H2a/b
Histone deacetylases interact with other proteins in order to achieve proper gene regulation control. Interacting
proteins can be for example transcription factors, methyltransferases and protein kinases and phosphatases.
Knowing these interacting proteins will point towards biological processes regulated by histone deacetylases and
hence open up new approaches towards their biological role. The project will identify and test novel proteins
interacting with histone deacetylases.
The aims of this project are:
a) To identify proteins interacting with histone deacetylases HD2a/b using a yeast two hybrid screen
b) To localise in living plant cells on the cellular level the interaction of HD2a/b with the identified
proteins using Bimolecular Fluorescence Complementation
c) To identify domains in HD2a/b required for protein interactions by testing HD2a/b mutant forms for
interaction with identified proteins.
3. Investigating the biological role of nitrilase interaction with 14-3-3 proteins during plant cell death
Plant nitrilases are enzymes thought to be involved in cyanide detoxification and hormone biosynthesis (2).
However, their true function is still under debate. My lab has shown that the nitrilases 1 to 4 interact with 14-3-
3 proteins. This indicates that the biological activities of these nitrilases are regulated by 14-3-3 proteins. This
regulation will be explored during the proposed honours project. We have further shown that induction of cell
death causes nitrilase 1: 14-3-3 complexes to localise to ER derived vesicles (figure 2b). The project will thus
explore the reason for this relocalisation and whether any of the other three nitrilases also localises to ER
derived vesicles during plant cell death. Finding these answers will help to understand similarities and
differences between the functions of the four nitrilase isoforms and will help us to understand their biological
roles.
The aims of this project are:
a) To investigate if 14-3-3 complexes with nitrilases 2, 3 and 4 localise to vesicular structures within the
cell during plant cell death.
b) To generate nitrilase proteins unable to bind to 14-3-3 proteins and to verify the loss of binding
c) To investigate whether loss of 14-3-3 binding changes the ability of nitrilases to localise to ER bodies.
References
(1.) Sridha and Wu, The Plant Journal 2006, 46, 124-133
(2.) Piotrowski 2008, Phytochemistry 69, 2655–2667
(3.) Comparot et al., 2003, Journal of Experimental Botany 54, 595-604
(4.) Bhat R.A. et al., Plant Methods 2006, 2:12
(5.) Cutler and Somerville, 2005, BMC Plant Biology, 5, 4
30
PROFESSOR ALLAN McKINLEY Room 2.11, Bayliss Building, Phone: 6488 3165
Email: [email protected]
My research interests involve: applications of spectroscopy for the detection and characterization of reactive
intermediates, theoretical modelling of the bonding in radicals, analysis and remediation of contaminated
groundwater, and biological applications of Electron Spin Resonance spectroscopy.
PROJECTS
1. Matrix isolation studies of reactive intermediates
We have built a state-of-the-art apparatus for measuring the ESR spectra of molecules trapped in solid neon at
4 K. There are less than half a dozen labs with this type of equipment in the world, no other in Australia. This
is cutting edge work and some of our recent successes CdCH3 [1], ZnCH3 [2], MgCH3 [3], Al2- [4], HgCH3 [5]
MgP, CdP and ZnP [6] are published in top international chemistry journals. We have also completed the
experimental phase for, MgN, ZnN, MgCH2 and MgCH radicals and articles on these molecules are in
preparation. The results of our studies are important to improve understanding of models of chemical bonding
as well as the chemical mechanisms involved in manufacturing computer chips, the wear-resistant coatings, and
even the chemical processes occurring in circumstellar dust clouds.
2. Radicals of Environmental or Astrochemical Relevance
We have been studying radical adducts formed between simples radicals such as OH, NH2 and O2 molecule and
a water molecule. To date we have published four papers in this area [7-10]. We are interested in nitrogen
containing radical adducts with water as these molecules could be important intermediates in the chemical
reactions occurring in our atmosphere or those of solar system bodies such as Titan, one of the moons of the
planet Saturn. The atmosphere of Titan is mainly nitrogen with traces of water and organic compounds. We are
also interested in the chemistry leading to the formation of methanol. Methanol has been observed on comets
and may be present on Titan. These experiments would involve matrix isolation IR and ESR experiments and
could involve PES experiments in collaboration with Professor Duncan Wild.
3. Environmental Chemistry of Contaminated Groundwater.
For some years now we have had a collaboration with Drs Greg Davis and Brad Patterson at the Land and Water
division of CSIRO at Floreat. In Australia, water is a key resource. In WA much of our water reserves are
underground and very vulnerable to pollution. We have studied the degradation in groundwater of BTEX
hydrocarbons (from leaking petrol stations), the mobility of pesticides such as atrazine and fenamiphos in soils
and we are evaluating the possibility of employing a new method for remediation of contaminated groundwater
using polymer-mats to introduce reagents into groundwater to promote microbial consumption of the pollutants.
As well as remediation of groundwater contaminated by BTEX and other volatile organics [12] we have studied
denitrification of ammonium nitrate contaminated groundwater[11]. There is a plume of ammonium nitrate
flowing into Cockburn Sound and we have tested this remediation technique on this plume [13]. In this field
study oxygen was introduced first to oxidize the ammonium ions to nitrate, and then ethanol was introduced
downstream to reduce the nitrate ions directly to nitrogen gas. Due to the scarcity of water there is also
considerable interest in ways of recycling and reusing water. Of particular interest is purifying waste-water
from sewage treatment plants with reverse osmosis equipment and using this water to recharge underground
aquifers. Questions that need to be answered include: how long do contaminants persist if they get through the
purification process and what chemical changes occur in the anoxic aquifer when oxygenated water is injected?
Projects in this area would involve either the analysis of the chemistry occurring in, or the mathematical
modeling of the mass transport phenomena involved with, pilot scale test-rigs for groundwater remediation
which are set up at CSIRO in Floreat.
31
4. Development of New Antimicrobials.
Multidrug-resistance in pathogenic strains of bacteria has in the last decade presented an increasing problem in
treatment of bacterial infections and diseases. The re-emergence of tuberculosis (TB), for instance, is one of the
serious threats and resistant strains of TB are rapidly spreading throughout the world. Furthermore, many strains
of enterococci have acquired resistance to vancomycin, one of the last lines of defence against such species.
Last year many wards at Royal Perth Hospital were plagued by VRE (vancomycin resistant enterococci) and
MRSA (methicillin resistant staphylococcus aureus) and a hospital in Melbourne reported the first cases of the
hypervirulent Quebec strain of Clostridium difficile.
In a joint project with Professors Riley (Microbiology) and Stewart (Chemistry) we have synthesized a new
compound which shows exceptional activity against gram-positive bacteria. The activity of this compound
against MRSA is similar to the activity of vancomycin and other commercial antimicrobials. We hold a
provisional patent on this compound and its analogues. Projects in this area could involve synthesis of
analogues of the compound with Professor Stewart or, for an appropriately qualified student, experiments with
Professor Riley to determine the mode-of-action of the compound and biological activity of its analogues.
References:
Copies are available from Dr Allan McKinley.
1.Karakyriakos, E.; Davis, J. R.; Wilson, C. J.; Yates, S. A.; McKinley, A. J.; Knight, L. B. Jr.; Babb R.; Tyler, D. J. ―Neon
and argon matrix ESR and theoretical studies of the 12CH3Cd, 12CD3Cd, 13CH3Cd, 12CH3111Cd, and 12CH3
113Cd Radicals‖ J.
Chem. Phys. 1999, 110, 3398-3410.
2.McKinley, A. J.; Karakyriakos, E.; Knight, L. B. Jr.; Babb, R.; Williams, A. ―Matrix isolation ESR studies of the various
isotopomers of the CH3Zn and ZnH radicals; comparisons with ab initio theoretical calculations‖ J. Phys. Chem. A 2000,
104, 3528-3536.
3.McKinley, A. J.; Karakyriakos, E. ―Neon matrix isolation ESR and theoretical studies of the various isotopomers of the
CH3Mg radical‖ J. Phys. Chem. A 2000, 104, 8872-881.
4.Stowe, A. C.; Kaup, J. G.; Knight, L. B. Jr.; Davis , J. R.; McKinley, A. J. ―Matrix-isolation investigation of the diatomic
anion radicals of aluminium and gallium (Al2- and Ga2
-): An electron resonance (ESR) and ab initio theoretical study.‖ J.
Chem. Phys. 2001, 115, 4632-4639
5.Karakyriakos, E.; McKinley, A. J. ―The Matrix Isolated HgCH3 Radical: An ESR Investigation‖ J. Phys. Chem. A. 2004,
108, 4619-4626.
6.Fuller, R. O; Chandler, G. S.; Davis, J. R.; McKinley, A. J. ―Matrix isolation ESR and theoretical studies of metal
phosphides‖, J. Chem. Phys. 2010, accepted for publication.
7.Langford, V. S.; McKinley, A. J.; Quickenden, T. I. "Identification of OH∙H2O in argon matrices." J. Am. Chem. Soc. 2000,
122, 12859-12863.
8.Cooper, P. D.; Kjaergaard, H. G.; McKinley, A. J.; Quickenden, T.I.; Schofield, D. P. "Infrared measurements and
calculations on H2O∙HO" J. Am. Chem. Soc. 2003, 125, 6048-6049.
9.Cooper, P. D.; Kjaergaard, H. G.; Langford, V. S.; McKinley, A. J.; Quickenden, T. I.; Robinson, T. W.; Schofield, D. P.
"Infrared Identification of Matrix Isolated H2O∙O2" J. Phys. Chem. A. 2005, 109, 4274-4279.
10.Ennis, C. P.; Lane, J. R.; Kjaergaard, H. G.; McKinley, A. J. ―Identification of the water amidogen radical complex.‖ J. Am.
Chem. Soc. 2009, 131, 1358-1359.
11.Patterson, B. M.; Grassi, M. E.; Davis, G. B.; Robertson, B.; McKinley, A. J. ―The use of polymer mats in series for
sequential reactive barrier remediation of ammonium-contaminated groundwater: laboratory column evaluation.‖ Environ.
Sci. Technol. 2002, 36, 3439-3445.
12.Patterson, B. M.; Davis, G. B.; McKinley, A. J. ―Polymer mats to remove selected VOCs, PAHs and pesticides from
groundwater: laboratory column experiments‖ Ground Water Monit. Rem. 2002, 22, 99-106.
13.Patterson, B. M.; Grassi, M. E.; Robertson, B. S.; Davis, G. B.; Smith, A. J.; McKinley, A. J.; ―The Use of Polymer Mats in
Series for Sequential Reactive Barrier Remediation of Ammonium-contaminated Groundwater: Field Evaluation.‖ Environ.
Sci. Technol. 2004, 38, 6846-6854.
32
WINTHROP PROFESSOR
HARVEY MILLAR ARC Centre of Excellence in Plant Energy Biology (PEB)
UWA Centre for Comparative Analysis of Biomolecular Networks (CABiN)
(www.plantenergy.uwa.edu.au, www.cabin.uwa.edu.au)
Bayliss Building, Room 4.74, Phone: 6488 7245
Email: [email protected] U
Cellular processes are directed by genes, orchestrated by proteins and delivered through fluxes of metabolites.
Using a combination of protein separation techniques, mass spectrometry and informatics my research group is
seeking to understand the compartmentation of cellular functions in cellular organelles and the networks of
molecules that define cell energy metabolism and its impact on real-world problems.
To see the latest publications from our group see:
http://www.plantenergy.uwa.edu.au/publications/millar.shtml
http://www.cabin.uwa.edu.au/publications
1. Senescence: remobilisation for plant productivity and yield.
With Dr Julia Grassl
The aging and dying of plant tissues (termed senescence) is an integrated and
essential process in plant development and has a critical role in remobilisation of
nutrients from leaves to both seeds and storage tissues. During this process nutrients
are transported from the outer leaf areas to the central vascular systems that feed the
growing plant. This can be seen as leaves turn colour in autumn. Re-localisation of
proteins and other molecules in this process is a large and important research area in
cereal crops. Finding molecular markers and genes that influence the senescence process could lead to plants
that perform better even in challenging environments such as nutrient deficient soils or during drought. A
project would use techniques like quantitative proteomic using isobaric labelling of proteins, molecular imaging
using mass spectrometry, Western Blotting, 2D gel electrophoresis, and transcript analysis.
2. Characterization of plant specific complex II subunits
With Dr Shoabai Huang
The mitochondrion is the powerhouse of the eukaryote cell by synthesis of ATP via
electron transport chain complexes coupled with the tricarboxylic acid (TCA) cycle.
Complex II (succinate dehydrogenase; SDH) has a central role in mitochondrial
metabolism as a component of both the electron transport chain and the TCA cycle.
Complex II catalyses the oxidation of succinate to fumarate. We have recently
shown that beyond its role in respiration, this protein complex is also involved in
defense signalling in plants by helping plants to respond to invading organisms like pathogenic fungi. The
objective of this project is to use T-DNA knockout lines of plant specific complex II subunits to characterise
their functions at the physiological, proteomic and metabolomic levels and therefore to uncover the hidden role
of these plant specific subunits .
3. Plant Mitochondrial Responses to Thermal Variation.
With Dr Nicolas Taylor
Fluctuations in temperature affect the metabolic processes of photosynthesis and
respiration and can have dramatic implications for biosynthesis, cellular
maintenance and growth. This can be seen in the different ways plants grow at low
and high temperatures. In this project you will be preparing cold and hot stressed
Arabidopsis plants and the isolating mitochondrial proteins from these plants. You
will then analyse these proteins by a quantitative proteomic technique using cutting
edge Q-ToF mass spectrometry. You will also analysis the mass spectrometry data to determine changes in
respiratory components in response to thermal variation.
4. Distinguishing Wheat Cultivars Using Mass Spectrometry. With Dr Nicolas Taylor Wheat flour is highly valued for its taste and dough-making properties. Because
these traits differ between cultivars, there is a need to readily identify cereal grains
according to cultivar especially with the development of cultivars that have
endpoint royalties, genetic modification or specific “built in attributes” that provide
agronomic or processing advantages. Currently it is almost impossible to distinguish
33
between cultivars from a seed sample prior to sowing or of a grain sample after harvest. This project aims to
determine novel biomarkers for commercial West Australian wheat varieties and develop these markers to allow
the distinction between these varieties. It will develop high throughput selective reaction monitoring (SRM)
assays to distinguish between wheat varieties. Your role in this project will be the preparation of protein extracts
from a range of commercial West Australian wheat varieties, analyse these samples using Q-ToF mass
spectrometry and collect proteins identifications for each cultivar. The differences in the proteins found for each
cultivar will be then used as biomarkers for each variety and SRM development
5. Proteomics of Rice phosphate stress-induced changes With Dr Ralitza Alexova
Phosphate is an essential element in a wide range of cellular components such as
macromolecular structures like nucleic acids, membrane lipids and proteins as well
as simple molecules like ATP and sugar phosphates. As most of the phosphate is
found in soils and rock deposits, organisms including plants have developed
strategies to extract this element and efficiently incorporate it into organic
molecules. This project will use high-throughput proteomic techniques to build a more complete picture of
global protein expression changes that occur in phosphate-stressed and phosphate-replete rice seedlings. The
rice phosphate stress response will be further investigated by developing novel mass spectrometry-based assays
for the simultaneous detection and quantitation of multiple proteins without the need for antibodies or chemical
labelling.
6. What determines nitrogen use efficiency in crop plants?
With Dr Clark Nelson
As nitrogen is the most expensive component in fertilizer production this nutrient is
arguably the most important component of plant metabolism to study. We are
exploring the biochemical machinery involved in nitrogen-use efficiency. In
collaborations we are conducting greenhouse trials as well as field trials in wheat,
barley, and rice to study the effects of various nitrogen regimes on metabolism. We
are applying a discovery-based approach using stable-isotope labelling and LC-MS techniques to monitor the
steady state proteome, and alteration in the metabolome of these plants. In this project you would be involved in
metabolomic and proteomic analysis of these cereal grains in an attempt to dissect the molecular mechanisms of
nitrogen metabolism.
7. Glutaredoxins as agents of redox homeostasis in plants
With Dr Elke Stroher
Posttranslational modifications (PTMs) of proteins, like formation of disulfide
bonds or addition of glutathione (glutathionylation), are important for fast
adjustment of protein activities – they can even serve as on/off switch for protein
activity. Members of the thioredoxin superfamily, such as thioredoxins (Trxs) and
glutaredoxins (Grxs) are the most likely candidates for re-reduction of oxidatively
modified proteins and are considered „key players‟ in signalling networks. This
project would consider Grxs in energy metabolism. Genetically modified Arabidopsis thaliana plants with either
less or more Grx would be analysed using novel biochemical and genetic technologies to uncover the impact of
this protein family in energy metabolism.
8. Biology of honeybee defence and reproduction
With Assoc/Prof Boris Baer and Dr Reza Zareie
Honeybees contribute to our economy and food industry by producing honey and
more importantly by pollinating some of our major crops and fruit trees. Honeybee
populations, however, have been declining in recent years, creating growing
concerns amongst many farmers and the public. To safeguard bees, research into the
bee immune system as well as their reproduction has been intensified on a global
scale. We have recently found that proteins within the honeybee seminal fluid
significantly increase sperm‘s life span. The next logical step is to isolate and identify which proteins are
responsible. In this project you will fractionate the seminal fluid and test which fractions are responsible to keep
sperm viable. We will then use mass spectrometry and other proteomics techniques to identify the proteins
behind this activity.
34
DR MATTHEW PIGGOTT ASSOCIATE PROFESSOR
Room 3.29, Bayliss Building, Phone: 6488 3170
Email: [email protected]
Synthetic Organic Chemistry, Medicinal Chemistry and Chemical Biology Our expertise in organic and medicinal chemistry is applied to the design and synthesis of therapeutic drug
candidates and small molecule probes to help investigate complex biological systems. We have several active
collaborations with more biologically orientated scientists and opportunities for cross-disciplinary projects exist.
The synthesis of biologically active natural products and novel aromatic molecules with potential applications in
organic electronics, supramolecular chemistry, and as components of molecular machines are other areas of
interest.
PROJECTS
1. Drug discovery for human African Trypanosomiasis
Human African Trypanosomiasis (HAT), also known as Sleeping Sickness, is caused by subspecies of the
protozoan parasite Trypanosoma brucei, transmitted by the Tsetse fly. Current treatments for HAT are toxic,
have difficult administration regimes and limited effectiveness, so there is a considerable need to find better
drugs. A recent high-throughput screen of the WEHI (Walter and Eliza Hall Medical Institute, Melbourne)
chemical library unearthed several promising hits, including the thiazole WEHI-1203394. Preliminary medicinal
chemistry in the Piggott group has identified the benzamide analogue MRK8 as having improved potency
against the parasite in vitro. This project will involve an expansion of this medicinal chemistry project in the
search for sub-nanomolar IC50 inhibitors of T. brucei.
NH
OS
NNH
OS
N
F
WEHI-12033940.48 M
MRK80.25 M
2. Chemical biology of phosphohistidines
with Professor Paul Attwood
Histidine kinases are a family of enzymes that catalyse the phosphorylation of the N1- or N3-imidazole nitrogen
of specific histidine residues in proteins. Their better-known cousins, the serine, threonine and tyrosine kinases,
have been implicated in the regulation of almost all eukaryotic cellular functions. In prokaryotes and lower
eukaryotes, histidine kinases play critical roles in the response to environmental stimuli. It is assumed that
histidine kinases and their substrates are also important components of mammalian cell-signalling pathways; for
example, Histone H4-kinase is upregulated in foetal, regenerating, and cancerous liver cells. However, none of
the mammalian histidine kinases are well characterised and their exact roles remain to be elucidated.
HN
NH
O
proteinprotein
NN
P
O
O
O
1
3
HN
NH
O
proteinprotein
HNN 1
3
histidinekinase
ATP
histidine residue N1-phosphohistidineresidue
H3N
N
O
O
NN
H2N
N
O
O
NN
POO
OPO
OO
13
1
3
HN
NH
O
proteinprotein
N
N3-phosphohistidineresidue
N
POO
O 3
OR 1
stable triazole analogue stable triazole analogue
35
The N-P bond in phosphohistidines is hydrolytically labile, which makes their identification, purification and
study challenging. For this reason, there are no antibodies to the phosphohistidine epitope, which impedes
progress in the field. We have recently devised syntheses of stable phosphonotriazole analogues of both isomers
of phosphohistidine. This project will involve efforts to exploit these compounds to learn more about
phosphohistidine biochemistry. Goals include the generation and characterisation of generic phosphohistidine
antibodies, affinity chromatography to purify histidine kinases and phosphohistidine-recognizing proteins, and
investigating the biological activity of the phosphonotriazoles as inhibitors of histidine kinases. This project
requires a combination of synthetic chemistry and biochemistry skills, but can be tailored to suit the strengths
and interests of the student.
3. Novel aromatic
molecular architecture
The classes of compounds
shown on the right are
challenging and
fundamentally interesting
synthetic targets, but also
have potential applications
in organic electronics,
supramolecular chemistry
and crystal engineering, and
as components of molecular
machines. Opportunities to
examine the metal
coordination chemistry and
electronic applications
(OFETs, OPVs and OLEDs)
of these compounds (once
synthesised) are possible
through collaboration with
Professor George
Koutsantonis and Dr Gavin
Collis (CSIRO Materials
Science and Engineering
Division, Melbourne).
4. A dihydropyrene-based organometallic molecular switch
with Professor George Koutsantonis
Dimethyldihydropyrenes are fascinating molecules with a planar 14- -electron periphery, making them
aromatic. They are easily converted to their valence tautomers, cyclophanedienes, by irradiation with certain
wavelengths of light. This project will involve the synthesis of a novel diethynyl-substituted dihydropyrene that
will be used to prepare organometallic complexes. Switching between the valence tautomers is expected to
drastically change the conductivity of the organic ligand, which in turn will affect properties such as colour,
crystallinity and redox potential.
5. Total synthesis of biologically active naphtho[2,3-
c]furan natural products
We recently achieved an efficient synthesis of the natural
product monosporascone. This project will use this
starting material for the synthesis of a number of related
secondary metabolites, including the antifungal agent
dehydroxyarthrinone.
O
O
O
OH
MeO
monosporascone(MAO inhibitor)
O
O
OH
OH
MeO
dehydroxyarthrinone(antifungal)
O
36
WINTHROP PROFESSOR
COLIN RASTON Founding Director, Centre for Strategic Nano-Fabrication (Incorporating Toxicology) and
Fledgling Centre for Green Chemistry and Molecular Discovery
Room 3.09, Bayliss Building, Phone: 6488 3045
Email: [email protected]
http://www.strategicnano.uwa.edu.au/
Organic Synthesis, Tissue Engineering, Nano-chemistry, Graphene, Desalination, Solar
and Fuel Cell Technology, Chemical Sensors, Drug Delivery, Microfluidics platforms
Current research covers: (i) Process intensification using spinning disc/rotating tube, electrospinning and narrow
channel processing, fabrication of nano-materials, nano-chemistry, supramolecular chemistry, and crystal
engineering, with applications in tissue engineering. (ii) Benign process technology – process intensification in
organic synthesis (controlling chemical reactivity and selectivity), and drug delivery. (iii) Device technology –
sensors, desalination, solar and fuel cells. Integration of these areas has led to novel chemistry and applications.
Projects for 2012 deal with these areas which are directed towards the major challenges facing humanity in the
21st Century – in being able to gain access to complex functional molecules and materials for tackling energy,
health and environmental issues. The projects are excellent training in a wide range of techniques, including
green chemistry, engineering, nano-technology, inorganic and organic synthesis, X-ray diffraction, NMR,
electron and atomic force microscopy, analytical techniques, other characterisation techniques. Brief details of
some projects are given below. Other projects are also available depending on the interests of the researcher.
PROJECTS
1. Controlling chemical reactivity and regio-selectivity in organic synthesis using microfluidic platforms
(MP) with Dr Keith Stubbs We have established the remarkable utility of MP in preparing organic compounds,
and projects here will focus on further applications in organic synthesis targeting molecules with biological
activity. There are two noteworthy effects of MP:
(i) Plug flow conditions which control chemo-selectivity without the need for
protection and de-protection.
(ii) The ability to control the kinetic and thermodynamic outcome of chemical
reactions which is not possible using classical stirred flask reaction vessels.
All this is under continuous flow conditions. In consequence of these findings we are
mapping out the plethora of organic reactions to establish the versatility of MP in
organic synthesis in general, and then to use the technology to prepare molecules with
particular function for biological applications. In the first instance we used MP to
prepare new classes of pyridine compounds which have application in medicine,
including diabetes inhibitors, and anti-cancer and anti-inflammatory activity, having
identified the binding prowess of molecules to G-Quadruplex (insert).
2. ‘Bottom up’ materials synthesis using dynamic thin films in microfluidic
platforms (MP) with Dr Paul Eggers and Dr Selvi Dev. We have recently
established that spinning disc processing (SDP) and rotating tube processing (RTP)
can be used to prepare nano-particles in a controlled way, for silver nano-particles
(medical and chemical catalysis applications), magnetite (medical imaging), gold
(medical technology), and drug molecules (drug delivery), and more. Recently we
patented a variable angle RTP which allows control over residence time, and is a
powerful MP for controlling the formation different shapes and nano-arrays. New
materials have potential in synthesis (eg. Heck reaction), medicine (e.g. multi-
functional imaging and drug release), and fuel cell and solar cell technology. The
MP facility at UWA is one of a few such facilities in the world, which is further
enhanced with a larger SDP and with accessible temperatures ≥ 600oC. Au, Pd and
Pt work is supported by The Perth Mint, with other projects in collaboration with
industry, eg drug delivery (with iCeutica), and tissue engineering (with Chimere
Pearls). Combining SDP with RTP, and narrow channel processors has exciting
possibilities in building complex functional nano-materials under continuous flow.
PIRS Nano-
Materials
PIRS
37
3. Applications of phosphonated calixarenes in tissue engineering and as
anti-cancer targets with Dr Keith Stubbs, Prof Fiona Wood, Prof Sarah
Dunlop, and Prof Lee Yong Lim. Relatively unexplored phosphonated
calixarenes have been prepared, 1, Fig 1, allowing access to derivatives with
alkyl chains attached to the phenolic O-centres, and various functionalised
moieties in the same position, including unsaturated chains (for photolytic
cross linking) and groups with specific binding prowess (towards metal
centres and organic molecules). Long alkyl chain (R) derivatives assemble
into intertwining nano-fibres with the overall material having nano-textured
features suitable for application in tissue engineering - for skin regeneration
and neurotrauma. In addition, the calixarenes can act as surfactants in
stabilising nano-particles (for imaging / magnetic field manipulation), and
binding drug and enzyme molecules, and as anti-cancer agents themselves.
4. New device technology for sensors, desalination and energy with Dr Swaminathan Iyer, Dr Paul Eggers
and Dr Ela Eroglu We recently developed drop casting devise technology for detecting hydrogen gas and
discriminating organic molecules in the gas phase. This has exciting possibilities in sensor technology for
detecting chemical warfare agents, fuel cells (including hydrogen gas), forensics (explosives and their
breakdown products) and solar cell technology. The core of the device is based on (i) carbon nano-tubes (CNT)
which can be decorated with selected nano-materials to tailor specific applications, and (ii) bare Pd nano-
particles, which are accessible using our recent advances in continuous flow technology microfluidics. Also
included is the development of new device technology for desalination.
5. Application of diatoms in device technology with Dr Swaminathan Iyer and Dr Ela Eroglu Single cell
diatoms have well ordered silica skeletons with regularly spaced pores all the same size with diameter di-
mensions down to 40 nm. The skeletons have exciting potential in nano-technology, ranging from medical (drug
delivery) through to solar and fuel cell technology, paint additives, water purification, and photonic devices.
We recently established that the pores can be decorated with nano-
particles of gold (inset), with a very narrow size distribution. The
proposed research focuses on extending it to decorating with
superparamagnetic nano-particles, associated with advances in the above
microfluidic platforms, as well as with several materials (different nano-
particles) depending on the application. High temperature treatment of
the skeletons is also possible using the new spinning disc reactor,
>600oC. This has potential in replacing silicon atoms with other metals,
titanium and magnesium, under scalable continuous flow conditions.
6. Materials chemistry of carbon with Prof Hui Tong Chua New forms of
carbon nano-materials, including composites of fullerenes with carbon nano-
tubes and graphene (as a recently established form of carbon), will be
investigated using self-assembly strategies and innovative approaches such
as high temperature continuous flow and scalable spinning disc processing.
A detailed understanding of the structures of these is important in developing
their potential applications. These include separating different diameter
carbon nano-tubes with different properties (semi-conducting versus
conducting), quantum dots, controlling chemical reactivity and selectivity
inside the tubes, chip devices for gas sensing (including chemical warfare
agents), devices for solar energy conversion, and desalination. Membranes
based on specific diameter carbon nano-tubes, in combination with other
material, will be developed to gain access to material for only water passing
across the membrane (desalination).
7. „Top-down‟ materials synthesis using microfluidic platforms (MP) We have recently established variable
angle rotating tube (VARTP) MP can be used to exfoliate graphite to graphene in
water, as a benign process, and similarly for boron nitride (BN). Varying the
conditions can result in the graphene and boron nitride sheets being „rolled up‟ into
scrolls, which have applications in chemical doping, hydrogen storage, battery
technology, and nano-mechanical devices. The intense shearing in the VARTP is
responsible for the „scrolling‟, and it has potential for exfoliating and/or scrolling
other laminar structures including mica and clays. The same shearing is also effective
in removing DNA from virus molecules, as an entry to drug delivery and vaccination
strategies using the resulting intact capsids, and also in controlling protein folding.
38
Dr. SAM SAUNDERS Room 3.10, Bayliss building, Phone: 6488 3153,
Email: [email protected]
ACER – Atmospheric and Environmental Chemistry
Research Group
My research interests have wide environmental implications. One of my keen interests is to measure
anthropogenic impacts, to develop practical tools for environmental impact assessment.
PROJECTS
Atmospheric Science and Air Quality Issues
1. Investigating reactive indoor air chemistry
This project will work towards extending the field of knowledge on indoor air chemistry. Particularly in
reference to the types of photochemical degradation reactions of organic compounds that occur in the indoor
environment and how these compare with those outdoor, for which there is currently very little research. The
project will focus on tailoring the Perth ambient outdoor model to the indoor study region and work on further
developing a new model for the simulation of the reactive indoor air chemistry based on the master chemical
mechanism (MCM) framework.
2. Assessment of the Pearl River Delta emissions, monitoring and meteorological data to develop a
regional chemical mechanism for simulating observed ozone formation and ambient VOC
measurements
Collaborating with the Hong Kong EPD, and Hong Kong Polytechnic University this project gives the
opportunity to make a significant contribution in developing a comprehensive tropospheric chemical
degradation mechanism, to provide simulations of the extensive sets of VOC and ozone measurements from
field campaigns in the Pearl River Delta region of China. A base case model has been developed in 2009, and
this requires further refinements for this geographic location. Only through developing an understanding of the
chemistry occurring in this airshed, will it be possible to work towards viable remediation strategies and reduce
the air pollution episodes in the region.
3. Simulating the experimental data from the CSIRO smog chamber facility
An important area in the development of air quality policy is in the use of experimental data from smog
chambers. Simulation of the experiments is used to help validate reaction mechanisms used in air quality
assessment. For this project data from several experiments conducted at the CSIRO smog chamber facility are
available, and there would be the possibility of a study visit to the facility at Lucas Heights in NSW. The project
aims to accurately simulate the experimental data, and investigate the impact of mechanistic details. Initial work
has focussed on 3 different VOC (toluene, 1,3-butadiene and isoprene) under different [VOC] and [NOx]
experimental conditions.
Other similar project developments may also be available in 2012, depending on collaborating partners.
And note projects will only be available for semester 2 in 2012.
39
All projects require an interest in gas phase chemical kinetics, reaction mechanisms and computational
chemistry, and to develop reaction schemes for volatile organic compounds. Background on the technology and
methodologies involved can be found on the following web site.
http://mcm.leeds.ac.uk/MCM/
Recent related publications
H. R. Cheng, H. Guo, S. M. Saunders, S. H. M. Lam, F. Jiang, X. M. Wang, T.J.Wang Photochemical ozone
formation in the Pearl River Delta assessed by a photochemical trajectory model Atmospheric Environment 44, 4199 (2010)
H. R. Cheng, H. Guo, X. M. Wang, S. M. Saunders, S. H. M Lam, F. Jiang, T. J. Wang, S. C. Lee, K. F. Ho –
On the relationship between ozone and its precursors in the Pearl River Delta: Application of an Observation-
Based Model (OBM). Environmental Science and Pollution Research 17:547–560 (2010)
S. Ho Man Lam , H. Cheng , H. Guo, S.M. Saunders X. Wang, I.J. Simpson, A. Ding, T. Wang, D. R. Blake.
A tailored Master Chemical Mechnism (MCM) model for the Pearl River Delta (PRD) region of South China.
19th
International clean air and environment conference, CASANZ, Perth, Sept. (2009) ISBN: 978-0-9806045
R.G. Hynes, D.E. Angove, S.M. Saunders, V. Haverd, M. Azzi – Evaluation of two MCM v3.1 alkene chamber
mechanisms using indoor environmental chamber data. Atmospheric Environment 39, p7251-7262 (2005)
S. Maisey, S.M. Saunders, N. West, P.J. Franklin.
Modelling Seasonal influences on Reactive Indoor Air
Pollution Chemistry for Residential Environs in the Southern Hemisphere. 19th
International Congress on
Modelling and Simulation, Perth, December (2011) http://mssanz.org.au/modsim2011
S.Maisey, P.Franklin, N.West, S.M. Saunders. Assessment of the indoor/outdoor relationship of VOCs in
residential properties in Perth, Western Australia. 19th
International clean air and environment conference,
CASANZ, Perth, Sept. (2009) ISBN: 978-0-9806045-1-1
40
PROFESSOR IAN SMALL ARC Centre of Excellence in Plant Energy Biology
Room 4.03, Bayliss building, Phone: 6488 4499
Email: [email protected]
Organelle Gene Expression Group
Our group is studying the RNA world within the energy organelles of plants – the mitochondria and chloroplasts.
These organelles contain the genes that code for the most important and abundant proteins on Earth, those that
drive photosynthesis, the basis for most biological productivity. The regulation of the expression of these genes is
crucial, yet still only poorly understood mechanistically. Our aim is to understand how the biogenesis and function
of chloroplasts and mitochondria are controlled through alterations in gene expression, with the goal of making
discoveries relevant to optimal use of plants in agricultural and environmental applications.
Gene regulation in plant organelles primarily occurs through changes in RNA processing, which makes these
expression systems unique. Much of our research builds upon the discovery of the PPR protein family, novel
sequence-specific RNA-binding proteins found in all eukaryotes, but particularly prevalent in plants (Schmitz-
Linneweber and Small, Trends Plant Sci, 13, 663-670). The experiments mostly involve the model plant
Arabidopsis thaliana to make use of the full range of international collections and databases on the ‗lab rat‘ of the
plant kingdom.
Prospective Honours students with a background in Molecular Biology, Biochemistry, Genetics or Computer
Science are particularly encouraged to apply. The projects will benefit from all the expertise and facilities
available within the ARC Centre of Excellence and will be at the forefront of research in this field.
PROJECTS
1. Analysing RNA processing with single-base precision by deep sequencing
RNA-seq is revolutionising the study of the transcriptome, by providing unprecedented detail into the nature of every
transcript in the cell. Although many RNA-seq studies limit themselves to simple quantitative measures of overall
gene expression, through careful design of experiments it is now possible to quantitatively analyse every step of
RNA processing (transcription, end-processing, splicing, polynucleotide tailing, editing…) in a single set of samples,
which would have been impossible only a year or two ago. The Centre‘s brand new HiSeq1000 is ideal for this
approach, and so there are a plethora of new possibilities. Some examples are listed below.
Mapping transcription start sites and processing sites. Primary transcripts differ from processed transcripts by
their 5‘ triphosphate; this leads to contrasts in the sensitivity of the RNAs to exonucleases and their ability to
be ligated. We can therefore distinguish these RNAs and use RNA-seq to map every transcription start and
processing site across the genome.
Analysing RNA editing. Plant organelle RNAs have their sequence changed after transcription, a mysterious
process referred to as RNA editing. These sequence alterations are highly specific and involve a particular set
of RNA-binding factors. Much remains to be discovered about RNA editing, and RNA-seq offers great
promise for analysing the process in more detail than ever before.
Identifying target sites for RNA binding proteins. Proteins bound to RNA can leave ‗footprints‘ by protecting
the RNA from exonuclease attack. By sequencing these footprints, we can discover where on the RNA the
protein was bound in the cell. RNA-seq allows us to do this across the entire transcriptome, mapping the
binding sites of multiple proteins at once. This information is crucial for working out how proteins recognise
their target sites (see ‗Deciphering the code‘, below).
Quantifying translation by ribosome footprinting. One set of particularly interesting ‗footprints‘ belongs to
ribosomes. By using antibiotics to block the ribosome in the course of translation, we can collect footprints
that show where the ribosomes were on each transcript. This gives unique information on the rate of
translation of each transcript, and insights into translational control mechanisms.
These projects will give students the chance to work with some of the most modern technology available anywhere.
It would suit students with a keen interest in biochemistry and/or genetics, especially those wanting to learn
computational data analysis approaches.
41
2. Deciphering the RNA-binding code of PPR proteins
in collaboration with Prof. Charlie Bond, Biochemistry
Studies of PPR proteins indicate that these proteins play key
roles in plant development, crop breeding and human
disease. In plants, the exact function of less than 10% of the
400-500 PPR proteins has been discovered. Several PPR
proteins have been shown to be essential for the expression
of genes required for the construction and function of the
major protein complexes involved in photosynthesis and
respiration. They are thus vital during germination and early
seedling development and some are absolutely required for
autotrophic growth. The current bottleneck in the study of
these important proteins is discovering the RNA targets of each one.
Statistical analysis of PPR protein sequences has suggested hypotheses proposing which amino acid residues are
required to recognise specific RNA targets. These hypotheses need to be tested experimentally by electrophoretic
mobility shift assays in which purified recombinant protein is incubated with labelled RNA target and run on
acrylamide gels - protein binding to the RNA oligonucleotide retards migration, giving a simple semi-
quantitative measure of binding affinity. It is simple in such assays to modify the sequence of the RNA, the
protein, or both, to investigate the importance of particular amino acids or particular bases in the target. These
experiments will lead to an understanding of which features in the RNA are being recognized by the protein, and
(we hope) to the ability to predict binding sites and even construct custom-designed proteins to bind desired
targets. The biotechnological possibilities are endless if we could achieve this. The project would suit students
interested in molecular biology, biochemistry or genetics.
3. Solving the mysteries of RNA editing in plant organelles
RNA editing is a site-specific modification of RNA molecules, occurring by nucleotide insertion/deletion,
nucleotide substitution or nucleotide modification. RNA editing alters the sequence of many different types of
RNAs in many organisms including plants and humans and constitutes a form of epigenetic gene regulation. In
many cases, RNA editing is essential for correct production of the protein encoded by the RNA, whilst in other
cases, RNA editing changes the functional properties of the encoded protein. In higher plants, RNA editing
consists of C to U changes and has been reported only in organelle transcripts, where over 500 different editing
sites are now known. Thirty PPR proteins have been found to be essential for the editing of specific sites in
organelle transcripts of Arabidopsis thaliana. These RNA-binding proteins probably constitute the specificity
factors recognizing the sequence around the target C. We have also identified a putative catalytic domain in
some PPR proteins that phylogenetically correlates with RNA editing.
We are approaching an understanding of how plants edit their organellar RNAs, with plausible hypotheses to
test, but experimental proof is lacking. There is an outstanding opportunity to for an Honours project to provide
the final data to confirm (or disprove!) the currently preferred model. The project will involve constructing
plasmids to express modified proteins in transgenic plants and then assaying for RNA editing. The project will
give a thorough grounding in many essential molecular biology techniques, including purification of DNA, RNA
and proteins; cloning, PCR amplification, bacterial and plant transformation, analyses of gene expression.
Relevant references from the group
Schmitz-Linneweber, C., and Small, I. (2008) Pentatricopeptide repeat proteins: a socket set for organelle gene
expression, Trends Plant Sci 13, 663-670.
Hammani, K., Okuda, K., Tanz, S. K., Chateigner-Boutin, A. L., Shikanai, T., and Small, I. (2009) A study of
new Arabidopsis chloroplast RNA editing mutants reveals general features of editing factors and their target
sites, Plant Cell 21, 3686-3699.
Chateigner-Boutin, A. L., and Small, I. (2010) Plant RNA editing, RNA Biol 7, 213-219.
Fujii S, Bond CS, Small ID (2011) Selection patterns on restorer-like genes reveal a conflict between nuclear and
mitochondrial genomes throughout angiosperm evolution. Proceedings of the National Academy of Sciences
USA 108(4):1723-8
42
PROFESSOR STEVE SMITH
ARC Centre of Excellence in Plant Energy Biology
and Centre of Excellence for Plant Metabolomics
Room 4.05, Bayliss Building, Phone: 6488 4403
Email: [email protected]
Discovering genes for plant growth
Arabidopsis thaliana provides the most powerful platform for modern genomics-based research in eukaryotes. It
provides us with the opportunity to discover genes and mechanisms by which plants grow, how they produce the
food that we eat, how they cope with environmental stresses (eg caused by climate change), and how they resist
diseases. Research using Arabidopsis can provide training in a range of disciplines including genomics, genetics,
cell biology, biochemistry, and new multi-disciplinary areas such as bioinformatics, systems biology and
metabolomics. The following projects are proposed but there is room for flexibility and originality, and the
emphasis can be matched to your particular skills or interests.
PROJECTS
1. Discovery of a new mechanism of growth control
Mutants that cannot break down their oil supply in the seed, fail to grow from seedlings into adult plants. But
wait! We have discovered two ways to persuade them to grow: 1) give them some sugar (ie. carbon, energy), or 2)
take away their supply of nitrogen (nitrate or ammonium)! This is very strange because it means that a seedling
deprived of carbon and nitrogen grows better than one that is deprived only of carbon! Our hypothesis is that the
seedlings „sense‟ and „measure‟ the relative amounts of carbon and nitrogen, and only grow when the ratio is
suitable. The original „oil mutant‟ is starved of carbon but has nitrogen. So either give it some carbon or take away
the nitrogen and it is happy.
Next, we have subjected our original „oil mutant‟ to mutagenesis and screened
the mutated progeny for seedlings that can now grow the same as wildtype (ie
with some nitrogen but without added sugar). There is one shown in the picture
among all its siblings that have not learnt the trick. This mutant should still be
unable to breakdown its oil supply, but is altered in its ability to „sense‟ or
„measure‟ the amounts of carbon and nitrogen. By identifying the new genes
which are mutated in such mutants we expect to discover molecular
components of the sensing or measuring pathway. In this way we should
identify new mechanisms of growth control in plants.
You will be given one or more mutants to study, with the aim of discovering
the mutated gene(s) and how it works. This will be an exciting journey of
discovery, taking us in an unknown direction. The project will likely involve
several techniques such as genetic analysis, molecular biology, metabolomics
and cell biology, and offers the potential to make important discoveries.
References
Germain V, Rylott EL, Larson TR, Sherson SM, Bechtold N, Carde JP, Bryce JH, Graham IA, Smith SM. (2001)
Requirement for 3-ketoacyl-CoA thiolase-2 in peroxisome development, fatty acid beta-oxidation and
breakdown of triacylglycerol in lipid bodies of Arabidopsis seedlings. Plant J. 28:1-12.
Martin T, Oswald O, Graham IA. (2002) Arabidopsis seedling growth, storage lipid mobilization, and
photosynthetic gene expression are regulated by carbon:nitrogen availability. Plant Physiol. 128:472-81.
43
2. Molecular mechanism by which karrikins from smoke promote seed germination
Karrikins are compounds discovered in smoke from bushfires, which promote seed
germination. They were discovered at UWA in a collaborative effort between Kings
Park botanists and UWA chemists. The original compound (structure 1, KAR1) is a
butenolide (3-methyl-2H-furo[2,3-c]pyran-2-one) but a few other closely related
compounds have also been found. We call them karrikins from „karrik‟, the first
recorded Noongar word for „smoke‟. It has been established that KAR1 can also
promote germination and seedling development in species that do not normally encounter smoke, raising the
possibility that karrikins represent a new class of plant growth-promoting substances of wide significance.
Karrikins have some structural similarity to a family of plant hormones called strigolactones, which can also
promote seed germination in some species, so they might act through a similar molecular mechanism.
Figure. Mutants that respond
differently to karrikins.
A. Karrikin insensitive (kai) mutant
(top row) does not germinate. The
„faker‟ (bottom row) is germinating
on KAR1 just like wildtype.
B. A genetic screen using a
transgenic line which is totally
dependent on KAR1 for germination.
C. Wildtype is inhibited by growth on
very high karrikin whereas a mutant
grows normally (okr „overdose on
karrikin resistant‟).
The goal of our research is to discover the molecular mode of action of karrikins in promoting seed germination
and seedling development. We are using Arabidopsis for this, both by studying existing mutants (eg. in seed
germination or hormone signaling) and by isolating new mutants. We have used transcript profiling with
microarray technology to identify genes that respond to KAR1. This provides insights into karrikin action as well
as a set of genes that can be targeted for mutation analysis. We have also carried out random mutagenesis of
wildtype Arabidopsis and isolated novel mutants that do not respond correctly to karrikins (see Figure). The aim
now is to discover the genes required for karrikin action and hence to discover its molecular mode of action. The
research will involve a range of techniques in molecular biology and biochemistry, and also close collaboration
with our chemistry friends.
References
Flematti GR, Ghisalberti EL, Dixon KW, Trengove RD. (2004). A compound from smoke that promotes seed
germination. Science. 305:977.
Nelson DC, Riseborough JA, Flematti GR, Stevens J, Ghisalberti EL, Dixon KW, Smith SM. (2009) Karrikins
discovered in smoke trigger Arabidopsis seed germination by a mechanism requiring gibberellic acid synthesis
and light. Plant Physiol. 149: 863-73.
Chiwocha SDS, Dixon KW, Flematti GR, Ghisalberti EL, Merritt DJ, Nelson DC, Riseborough JAM, Smith SM,
Stevens JC. (2009) Karrikins: A new family of plant growth regulators in smoke. Plant Science 177: 252-256.
Nelson DC, Flematti GR, Riseborough JA, Ghisalberti EL, Dixon KW, Smith SM (2010) Karrikins enhance
light responses during germination and seedling development in Arabidopsis thaliana. Proc. Natl. Acad. Sci.
USA, 107, 7095-7100.
44
PROFESSOR MARK SPACKMAN Room 4.11, Bayliss Building, Phone: 6488 3140
Email: [email protected]
Crystallography and theoretical chemistry Our research investigates in detail the structure of crystals, in particular the electron distribution and properties
related to it, such as electric moments of molecules (dipole, quadrupole, etc.), electrostatic potential and electric
field, and also measures of its response to external perturbations, including polarizability and hyperpolarizability.
All research projects in this area incorporate different aspects of physical and theoretical chemistry. They utilise
ab initio computational methods along with some computer programming and computer graphics and, where
applicable, measurement and detailed analysis of high-resolution, low-temperature X-ray diffraction data.
The Honours projects listed below will provide valuable practical experience with the techniques of modern
computational chemistry and a familiarity with state-of-the-art ab initio quantum chemical calculations, as well
as some practical experience in the use and applications of X-ray crystallography. The amount of hands-on
experience with computer programming and graphics on the one hand, and experimental measurement of X-ray
diffraction data on the other hand, can be tailored to suit the project and the candidate.
PROJECTS
1. Electrostatic complementarity as a guiding principle in molecular crystals
with A/Prof Dylan Jayatilaka and Dr Mike Turner
In recent years much of our research has focused on a
detailed exploration of the attributes and uses of
Hirshfeld surfaces, which are now making a substantial
contribution to the improved understanding of
intermolecular interactions in bulk materials, and
especially crystal engineering (the understanding of
intermolecular interactions in the context of crystal
packing and exploiting that understanding in the design
of new solids with desirable physical and chemical
properties). Details and examples of this exciting work,
including the program CrystalExplorer, developed in
collaboration with Dylan Jayatilaka's group, can be
found at the web site associated with this project:
http://ra.bcs.uwa.edu.au/CrystalExplorer/.
This Honours project will explore in more detail the
electrostatic potential mapped on these surfaces,
especially the way in which the electropositive part of one molecule coincides with the electronegative region of an
adjacent molecule (an example is given in the figure). This qualitative picture of intermolecular interactions will be
compared with the more quantitative results obtained with ab initio calculations of intermolecular interaction
energies, and for a range of molecular crystals incorporating hydrogen bonds, halogen bonds and other important
interactions.
45
2. Charge density analysis of fundamental host-guest supramolecular systems
several projects, with A/Prof George Koutsantonis and Dr Alex Sobolev
Although supramolecular chemistry is one of the most active fields of modern chemistry, very little seems to be
known about the detailed nature of the host and guest systems that comprise these aggregates. Supramolecular
systems – molecular aggregates – underpin the design and development of materials in areas as diverse as catalysis,
targeted drug delivery, gas storage, chemical separation and nonlinear optics. They also serve as models for
complex phenomena such as self-assembly and ligand-receptor binding. Projects in this area are part of a research
program aimed at a greater understanding of intermolecular interactions and the properties of host-guest systems in
the solid state, particularly organic clathrates and complexes formed by small molecules interacting with crown
ethers, calixarenes, molecular tweezers and cages (some examples are given in the figure below). These projects
will involve some synthesis, and measurement of highly accurate X-ray diffraction data, complementary neutron
diffraction experiments, quantum chemical calculations and computer graphics. A particular focus of the charge
density analyses will be the polarization and dipole moment of guest molecules as a function of the changing
electrostatic nature of the host systems.
3. Reactivity in crystals and its relationship to voids and cavities
with A/Prof Dylan Jayatilaka and Dr Mike Turner
Reactivity in crystals has been the focus of increased activity in recent years, in particular the recent kinetic studies
of E/Z photoisomerizations occurring in co-crystals, [2+2] photodimerizations in organic crystals (for example, (a)
to (b) in the adjacent figure) and single-crystal to single crystal transformations in molecular framework materials.
Many studies such as these use concepts of "reaction cavity"
and "void space" to rationalize the observed reaction products,
and in particular the differences between solution and solid
state products. The Hirshfeld surface (see Project 1, above) is a
measure of the space occupied by a molecule in a crystal, and
hence it should be able to provide a considerable amount of
relevant information, or at least a vehicle for mapping
properties such as the magnitude of the LUMO orbitals, etc.
This project will build on the results of Maram Susli, a 2009
Honours student, to further explore the correlation of void
locations, volumes and orbital properties with experimental
information on various kinds of reactivity involving molecules
in crystals.
Another aspect of this project could focus on a more detailed
investigation of void space (i.e. empty space) in reactive solids such as
metal-organic frameworks (MOFs) and zeolites. An example of the void
space in Linde type A zeolite is shown in the figure on the right. This
project would exploit the recent implementation of tools in
CrystalExplorer for visualising and mapping void surfaces and volumes.
46
DR SCOTT STEWART Room 3:30, Bayliss building, Phone: 6488 3180,
Email: [email protected]
Research Overview
Research interests include the construction of biologically active natural products utilising modern organic
synthetic methods. Many these syntheses are designed using palladium catalysed cross coupling reactions as the
key step transformation. Several natural products Arboflorine (1),1 Ajamalicine, Pumiliotoxin B (2) ,
2 Flinderole
A,3 Epoxiquinol and BE-26554B (3) have been targeted within this group because of their interesting molecular
architecture and biological activity. Related to this field, methodological studies involving the improvement
various reactions including, Suzuki, Buchwald-Hartwig and intramolecular Heck reactions through the
modification of nickel(0) and palladium(0) catalytic conditions are currently being explored. Research in the
discovery of novel domino transformations (the execution of two or more bond-forming transformations under
identical reaction conditions)4 mediated by palladium are routinely carried out within the group.
1b,5 Medicinal
chemistry interests include the synthesis of libraries of new thalidomide analogues for the inhibition of tumour
necrosis factor (TNF) expression as well as determining the molecular mode of action.6,7
1. The Synthesis of Ngouniensine and Arbiflorine
The domino Tsuji-Trost/Heck reaction has been used devised within our group and used for the construction of
the azepino[4,5-b]indole ring system 5 and 3-benzazepines. In this process the construction of the seven
membered C-ring can be achieved in a single step. This project will involve using this domino reaction as a key
step for the production of the natural product Ngouniensine (6). The natural product alkaloid 6 isolated from
Strychnos ngouniensis has reported activity against several P. falciparum strains, a protozoan parasite
responsible for the cause of malaria in humans. Although the IC50 value of 6 is moderate the epimer at C20 is
more potent suggesting that analogues generated at C20 should be investigated.
Although the domino Tsuji-Trost/Heck reaction is sufficient for the construction of the azepino[4,5-b]indole ring
system 5, the exocyclic olefin within this ring system is not amenable to use in the total synthesis of
Arboflorine.1 The second part of this project is to investigate reactions, namely reaction between tryptamine and
methyl chloropyruvate, in the production of unsaturated azepino[4,5-b]indoles and their use as precursors for
Arboflorine.
2. The Synthesis of Amphibian Alkaloids through the Tsuji-Trost Reaction
Several classes of compounds can be found in the skin of Amphibians with a wide range of biological activity.
One such class of compound includes the pumiliotoxins of the general indolizidine structre 8 where R is an alkyl
side chain. Several pumiliotoxins have cardiotonic and myotonic activity through binding to unique binding sites
47
on the voltage dependent ion channel. This project will focus on the synthesis of the indolizidine ring system
staring with proline 2 which includes the correct stereochemsistry at the -carbon. Previous group work has
generated the indolizidine core through a key intramolecular Tsuji-Trost cross coupling reaction.8 Such
palladium mediated cross coupling reactions have been used regularly in complex natural product synthesis.
3. Enantioselective and Diastereoselective Domino Reactions (with Dr F. Pfeffer)
In 2010, we reported a new domino reaction for the production of tetrahydro-β-carbolines 10 which involves an
initial Heck reaction followed by an aza-Michael addition.5a
This process, also amenable to many acrylate based
reagents resulting in variation of the terminal functional group, is considered a process comparable to the Pictet-
Spengler reaction. In this domino reaction, however the stereogenic centre at C* is not formed in a controlled
manner. The aim of this project is to use a chiral pool variation of the toluenesulfonyl (Ts) protecting group at
N10
to create a diastereoselective domino reaction firstly generating epimers at C*. A second part of this project
will involve the investigation of an enantioselective domino process by altering the using enantiopure
phosphines or quaternary ammonium salts. Once the generation of the stereocentre at C* is confirmed then an
application in the synthesis of biologically active natural products such as Elaeocarpidine 11 is to be attempted.
4. The Preparation of New Reagents and Catalytic Systems in Organic synthesis
Organocatalysis is a rapidly developing field in synthetic organic chemistry. In a simple organic reaction such as
the Michael reaction new stereogenic centres can be generated asymmetrically in a single step in high yields and
ee. The aim of this project is to create new organocatalytics reagents and reactions. In particular an asymmetric
variant of Mander‘s reagent 13 and/or applied transformations will be investigated. This reagent has been
effectively used for -keto ester formation in fine chemical and natural product syntheses.
References
1.a) Lim, K-H.; Kam, T-S. Org. Lett., 2006, 8, 1733; b) S. G. Stewart, C. H. Heath, E. L. Ghisalberti, Eur. J.
Org. Chem, 2009, 1934. 2. J. Daly, T. Spande and H. Garraffo, J. Nat. Prod., 2005, 68, 1556-1575. 3.
Fernandez, L. S., Buchanan, M. S.,et al., Org Lett 2009, 11 (2), 329-332 4. L. F. Tietze.; G. Brasche.; K. M.
Gericke, Domino Reactions in Organic Synthesis, Wiley-VCH 2006. 5a) D. L. Priebbenow, L. C. Henderson, F.
M. Pfeffer, S. G. Stewart, J. Org. Chem. 2010, 75, 1787; c) S. G. Stewart, E. L. Ghisalberti, B. W. Skelton, C. H.
Heath, Org. Biomol. Chem, 2010, 8, 3563. 6.a) S. G. Stewart, L. A. Ho, M. E. Polomska,
A. T. Percival, G. C. T.
Yeoh, ChemMedChem, 2009, 4, 1657; b) S. G. Stewart, M. E. Polomska, R. W. Lim, Tetrahedron Lett. 2007, 48,
2241. 7.a) S. G. Stewart.; D. Spagnolo, M. E. Polomska, M. Sin.; M. Karimi.; L. J. Abraham, Bioorg. Med. Chem
Letts. 2007, 17, 5819; b) S. G. Stewart.; C. Braun.; S-L. Ng.; M. E. Polomska.; M. Karimi.; L. J. Abraham,
Bioorg. Med. Chem. 2010, 18, 650; c) S. G. Stewart, C. J. Braun, M. E. Polomska, M. Karimi, L. J. Abraham, K.
A. Stubbs, Org. Biomol. Chem., 2010, 8, 4059; 8. R. E. Martin, M. E. Polomska, L. T. Byrne, S. G. Stewart
Tetrahedron Lett. 2011, 4878.
48
DR KEITH STUBBS Room 4.18/Lab 4.22, Bayliss Building, Phone: 6488 2725
Email: [email protected]
Research Interests
Carbohydrates are present in every living system from prokaryotes to eukaryotes and traditionally, have been
known for their role in the structural integrity of plants and as energy sources. Recently, however, carbohydrates
have been shown to be involved in a variety of fundamental biological processes such as protein folding and
trafficking, as well as cellular signaling and recognition. As we gain greater understanding into the roles that
carbohydrates play at the cellular level, scientists are faced with new challenges. On the chemistry side, unique
carbohydrate-based tools need to be developed and in turn used to investigate the specific roles that a single
mono- or polysaccharide plays in the dynamics of the cell in order to keep up with the biochemical discoveries
of new glycan structures and the enzymes that regulate them. My research aims are to address the development
of such tools.
The laboratory is a highly collaborative environment where researchers work to solve problems in chemical
glycobiology. Depending on the project, you will have the opportunity to gain exposure to methods ranging
from carbohydrate synthesis, protein expression, molecular biology and enzymology. The laboratory enjoys
extensive collaborations and researchers are provided with mentoring so as to aid their scientific development
and enable them to realize their professional goals. All the summaries of projects outlined below will initially
involve the synthesis of compounds and once prepared, investigation(s) using biochemical and microbiological
assays will be conducted.
If you are excited about interdisciplinary science, enjoy experimental research in chemistry or biochemistry and
are interested in joining the laboratory feel free to contact Dr. Stubbs by email or come and chat with me about
these and any other projects and research interests you are interested in.
PROJECTS
1. Development of new scaffolds to inhibit carbohydrate-processing enzymes.
The enzymes that regulate the structures of glycans are extremely
important and have been implicated in a wide variety of diseases and
thus are targets for therapeutics. For example, carbohydrate-
processing enzymes are important for bacterial growth and invasion
of our cells. Project(s) described here will be to design and
synthesize new inhibitor scaffolds that can be used to investigate the
role these carbohydrate-processing enzymes play in human disease.
The prepared compounds will be tested for their potency against the
human enzymes in question and they will also be tested in vitro to
determine their effectiveness at the cellular level. As well, through
strong international collaborations, these compounds will also be co-
crystallized with proteins of interest (example on left). This research
is funded by the Australian Research Council (ARC).
Students with interests in synthetic chemistry or both synthetic chemistry and biochemistry are very well suited
for this project.
2. Investigations into the glycobiology of Helicobacter pylori.
Helicobacter pylori is a Gram-negative, microaerophilic bacterium that infects the
stomach and duodenum. It has been shown that many cases of peptic ulcers, gastritis,
duodenitis, and stomach cancers are caused by H. pylori infections. Whilst a lot of
information has been gathered on the genetics and pathology of H. pylori infection, the
role that carbohydrates play in this bacterium‟s life cycle and in mediating host-pathogen
interactions is lagging. Increased insight into these interactions would be of use in the
design of new therapeutics to treat H. pylori infections.
49
In collaboration with Professor Alice Vrielink, Associate Professor Mohammed Benghezal and Professor Barry
Marshall, projects under this heading will investigate, through chemical synthesis and molecular biology, what
roles carbohydrates and larger glycan structures play in the pathogenesis of H. pylori and to use this information
in the design of new therapeutics. This research is funded by the National Health and Medical Research Council
(NHMRC).
Students with interests in synthetic chemistry or both synthetic chemistry and microbiology are very well suited
for this project.
3. Investigations into the glycobiology of Neisseria sp.
Neisseria sp. are Gram-negative bacteria that colonize the mucosal surfaces of many animals. Of interest are the
two pathogens Neisseria meningitidis, which causes bacterial meningitis, and Neisseria gonorrhoeae, which
causes gonorrhoea. These two pathogens have developed unique mechanisms of invading host cells many of
which involve carbohydrates and their associated enzymes. Insight into these interactions would be of use in the
design of new therapeutics to treat Neisseria sp. infections. In collaboration with Professor Charlene Kahler and
Winthrop Professor Alice Vrielink projects under this heading will investigate, through chemical synthesis and
molecular biology, what roles carbohydrates and larger glycan structures play in the pathogenesis of Neisseria
sp. This research is funded by the National Health and Medical Research Council (NHMRC).
Students with interests in synthetic chemistry are very well suited for this project.
50
Dr. K. Swaminathan Iyer ARC Australian Research Fellow
Deputy Director, Centre for Strategic Nano-Fabrication,
School of Biomedical Biomolecular and Chemical Sciences
Phone: 6488 4470, Bayliss, Room: 4.41.
Email: [email protected]
BioNanoChemistry: Interdisciplinary research encompassing Chemistry, Physics and
Biology.
PROJECTS
1. Magnetically responsive polymeric scaffolds for wound
healing with Prof. Fiona Wood, Prof. Tim St. Pierre, Dr. Rob
Woodward and Dr. Mark Fear. Despite recent therapeutic advances,
the mortality and morbidity from major burns remains high.
Consequently, there is a pressing need to develop economical,
efficient and widely-available therapeutic approaches to enhance the
rate of wound re-epithelialization and restoration of the protective
epithelial barrier. Skin, the largest organ of the human body, provides
an essential protective barrier and serves several homeostatic/sensory
functions vital to health and its functional recovery post burn injury
remains the ultimate goal of wound healing research. Polymer
nanoscaffolds have been extensively utilized in the design of tissue
engineered constructs in delivering several growth factors for the correction of a wide range of medical
conditions. A variety of polymeric scaffolds have been used to deliver growth factors, including natural or
synthetic polymers that generally form either hydrogels or solid polymer scaffolds. However extended release of
proteins is not easily achieved due to the release kinetics of growth factor through hydrogels being mainly
diffusion controlled via the numerous aqueous channels within the hydrogels. Immobilization of the growth
factor within the biodegradable hydrogel seems to improve the release kinetics, with release being controlled by
the degradation of the hydrogel. Here the release kinetics are slow and progressive necrosis sets in post injury. A
novel modulated delivery system would indeed be ideal, allowing the release profiles of payloads to be
manipulated to match the physiological requirements of the patient. The project will explore the utility of
magneto-responsive scaffolds for on-demand delivery of payloads.
2. Exploring nanoparticles as biomarkers in evolutionary biology with Dr. Boris Baer.
This project is collaboration with Collaborative Initiative for Bee Research (CIBER: http://www.ciber.science.uwa.edu.au/) and the BioNano Research Initiative in
Chemistry. Researchers in CIBER have been investigating honeybee reproduction,
which is quite spectacular, as queens only mate at the beginning of their life, during one
or very few mating flights. Following this they are able to store millions of sperm for
years, and use them in very economic ways to fertilize millions of eggs. Currently there
is very little information how social insect queens are able to keep sperm alive for years,
how active sperm remain during storage, and
how queens are able to economize their use
of sperm during egg fertilization. This
research project explores the possibility of
developing nanoparticles as markers in an
attempt to unravel this phenomenon by
tagging sperms. The project will involve
synthesis of magnetic nanoparticles and semiconductor quantum dots as markers,
exposure to the bee research team in CIBER and training at the nanotechnology
and biology interface.
3. Antibody conjugation of nanoparticles for cell specific drug
delivery in the central nervous systems with Dr. Lindy Fitzgerald and
Prof. Sarah Dunlop.
In the nervous system, all sensations and behaviors are encoded in dynamic patterns of activity in cellular
networks. Through a sequence of neural networks, sensory information is transmitted to higher, associative
brain areas. Following integration in these areas, specific activity patterns are eventually formed in the relevant
motoneuron pools to produce adequate behavior. In this chain of events, key processing steps are thought to
51
occur on the level of local microcircuits that contain on the order of 1,000–10,000 cells. These local circuits
form highly connected three-dimensional networks. Calcium ions (Ca2+) have been a favorite target in
molecular imaging studies because of the important role of calcium as a second messenger in cellular signaling
pathways. Neurotrauma, such as traumatic brain or spinal cord injury, encompasses both acute damage induced
by the primary injury and chronic progressive secondary degeneration of intact, but highly vulnerable, tissue,
results in a drastic change in the cellular signalling pathways. Reactive oxygen and nitrogen species (ROS and
RNS) are implicated to play a vital role in this, as their production is reported to exceed a cell‘s antioxidant
capacity following injury. After neurotrauma, calcium fluxes are uncontrolled and spread to intact but
vulnerable tissue. Triggers for uncontrolled calcium fluxes are varied but include ROS/RNS which activate
Ca2+ channels and repress Ca2+ pumps. Astrocytes in particular are chiefly responsible for the brain‘s
antioxidant defense whereby they play a pivotal role in protecting
neurons and oligodendrocytes from oxidative stress. The project explores
developing cell specific targeting of astrocytes to develop drug delivery
vehicles as a mode to combat secondary degeneration following
neurotrauma. This site specific targeting is projected to have high
efficacy regulation in the calcium flux.
4. Developing a nanoscale therapy to alleviate oxidative stress in
placental-related diseases of pregnancy with Prof. Jeff Keelan and
Prof. Brendan Waddell.
Pregnancy is a state of oxidative stress arising from increased placental
mitochondrial activity and production of reactive oxygen species (ROS),
mainly superoxide anion. The placenta also produces other ROS
including nitric oxide, carbon monoxide, and peroxynitrite which have
pronounced effects on placental function including trophoblast
proliferation and differentiation and vascular reactivity. Excessive
production of ROS may occur at certain windows in placental
development and in pathologic pregnancies, overpowering antioxidant
defences with deleterious outcome. For example: miscarriage and pre-
eclampsia are the most common disorders of human pregnancy. There is
mounting evidence that oxidative stress or an imbalance in the
oxidant/antioxidant activity in utero–placental tissues plays a pivotal role
in the development of placental-related diseases. This project explores
the application of magnetic nanoparticles as antixodant delivery agents in
placenta via a systematic approach.
5. Colloidal Upconverting NaYF4 Nanocrystals Doped with Er3+
,
Yb3+
and Tm3+
for biomedical imaging and diagnostics with Prof D. D. Sampson. Upconversion
nanocrystals are luminescent nanomaterials that convert a near-infrared excitation into a visible emission
through lanthanide doping. Compared to organic fluorophores and semiconducting nanocrystals, upconversion
nanocrystals offer high photochemical stability, sharp emission bandwidths, and large anti-Stokes shifts (up to
500 nm) that separate discrete emission peaks from the infrared excitation. Along with the remarkable light
penetration depth and the absence of autofluorescence in biological specimens under infrared excitation, these
upconversion nanocrystals are ideal for use as luminescent probes in biological labelling and imaging
technology. Organic dyes and semiconductor quantum dots that emit at higher energies via two-photon
absorption processes require expensive high energy pulse lasers. Due to the relative high efficiency of the
upconversion process in lanthanide-doped materials,
inexpensive 980 nm NIR diode lasers may be employed
as the excitation source. The realization of efficient
NIR to visible upconverting nanocrystals can be
exploited to develop novel dual modality drug carriers.
The project will explore the synthesis and properties of
doped NaYF4 nanosystems and their utility as
biomarkers in vitro. See for reference: Analyst, 2010, 135, 1839-1854.
Diagram of a gestational sac
at the end of the 2nd month
showing the myometrium (M),
the decidua (D), the placenta
(P), the exo-coelomic cavity
(ECC), the amniotic cavity
(AC) and the secondary yolk
sac (SYS). Ref: Human
Reproduction Update,
Volume12, Issue6, Pp. 747-
755.
52
6BASSOCIATE PROFESSOR
ROBERT TUCKEY 29BRoom 3.71, Bayliss Building, Phone: 6488 3040,
30B Email: [email protected] U
Molecular Steroidogenesis Group
Current research involves the metabolism of vitamins D2 and D3 by cytochrome P450scc, and the activation
and inactivation of vitamin D by other mitochondrial-type cytochromes P450 including CYP27A1, CYP27B1
and CYP24.
Mitochondrial Cytochrome P450 Enzymes There are 7 mitochondrial cytochrome P450 enzymes encoded by the human genome. They catalyse
hydroxylation reactions involved in steroid hormone synthesis and the metabolism of vitamin D. The
mitochondrial P450s receive electrons to support their hydroxylation reactions from NADPH via adrenodoxin
reductase and adrenodoxin. These P450s appear to be anchored to the mitochondrial membrane primarily by a
region involving the F-G loop and bind substrate from the hydrophobic domain of the inner-mitochondrial
membrane (Figure 1). Cytochrome P450scc (CYP11A1) catalyses the conversion of cholesterol to
pregnenolone, termed the cholesterol side-chain cleavage reaction. This reaction involves three hydroxylations,
all of which occur at a single active site on cytochrome P450scc. Pregnenolone serves as the precursor of all the
steroid hormones such as corticosteroids, androgens and estrogens.
In collaboration with Professor Andrzej Slominski at the University of Tennessee, Memphis, we tested the
ability of P450scc to metabolize vitamins D2 and D3. These potential substrates, structurally similar to
cholesterol, were incubated with purified P450scc and in some cases were also incubated with P450scc in rat
adrenal mitochondria. Products were purified by TLC or HPLC and identified by mass spectrometry and/or
NMR. We found that human and bovine P450scc did not cleave the side chain of vitamin D3 but hydroxylated
the side chain producing 20-hydroxyvitamin D3, 20,23-dihydroxyvitamin D3 and 17,20,23-trihydroxyvitamin
D3. P450scc converted vitamin D2 to 20-hydroxyvitamin D2 and 17,20-dihydroxvitamin D2, again with no
cleavage of the side chain occurring.
We have carried out biological testing of several of the novel P450scc-derived hydroxyvitamin D3 products in
collaboration with Professor Slominski in Memphis. 20-Hydroxyvitamin D3 has been found to be as effective as
the hormonally active form of vitamin D3, 1,25-dihydroxyvitamin D3, in inhibiting cell proliferation and
promoting differentiation of a variety of cells including skin, leukaemia, breast and prostate carcinomas.
Importantly, it does not raise serum calcium levels in rats and consequently lacks the toxic side effect of
hypercalcaemia caused by high doses of 1,25-dihydroxyvitamin D3. 20-Hydroxyvitamin D3 thus shows promise
as a therapeutic agent for the treatment of hyper-proliferative disorders including cancer.
Other mitochondrial P450s we are studying are CYP27A1, CYP27B1 and CYP24, all of which can act on
vitamin D. CYP27A1 catalyses the first step in the activation of vitamin D which is hydroxylation in the 25-
position. CYP27B1 catalyses the second step in the activation of vitamin D, 1α-hydroxylation of 25-
hydroxyvitamin D3 to produce 1,25-dihydroxyvitamin D3, the hormonally active form of vitamin D. 1,25-
Dihydroxyvitamin D3 not only regulates calcium metabolism, but also has many other important effects
including inhibiting proliferation and promoting differentiation of a range of cells, plus regulating the immune
system. CYP24 acts on 1,25-dihydroxyvitamin D3, hydroxylating it at C24 which causes its inactivation. We are
Figure 1. Model of the
interaction of cytochrome
P450scc with the inner-
mitochondrial membrane
53
expressing CYP27A1, CYP27B1 and CYP24 in E. coli, and studying their catalytic properties in a reconstituted
system that utilizes phospholipid vesicles to mimic the inner-mitochondrial membrane. We are also using these
enzymes to hydroxylate the P450scc-derived vitamin D analogues such as 20-hydroxyvitamin D3 to see if 1-,
24- or 25-hydroxylation of these compounds enhances their potency without returning their calcaemic activity,
with the aim of further improving their therapeutic potential.
Of the 7 mitochondrial P450s in humans, only one remains whose function is unknown, CYP27C1. Since this is
in the same family as CYP27A1 and CYP27B1, both of which can act on vitamin D derivatives, is likely that
CYP27C1 acts on a vitamin D analogue or a steroid with similar structure. Mitochondrial P450s are also found
in invertebrates but to date are poorly characterized. There are three mitochondrial P450s in insects that catalyse
steroid hydroxylations, similar to what occurs in mammals for the synthesis of active steroids from cholesterol.
In the case of insects the final active hormone is 20-hydroxyecdysone, also known as insect moulting hormone.
The final activating step in the synthesis of ecdysteroids is the addition of the 20-hydroxyl group by
mitochondrial CYP314A1.
PROJECTS
1. Can human CYP27C1 metabolize vitamin D or steroids?
CYP27C1 is the only mitochondrial P450 in humans whose function is yet to be determined. It has been
expressed in E. coli and purified but no substrate for this enzyme has yet been identified. The CYP27C1 is
expressed in a number of tissues including liver and kidney, known sites of vitamin D activation. The other two
members of the CYP27 family, CYP27A1 and CYP27B1, are known to metabolise vitamin D hydroxylating it
in the 25- and 1- positions, respectively. It would seem most likely that CYP27C1 will also act on a vitamin D
derivative, but the only ones tested to date are vitamin D, 25-hydroxyvitamin D and 1-hydroxyvitamin D, where
no metabolism was found. The aim of this project is to express human CYP27C1 in E. coli, purify the expressed
enzyme and examine its ability to hydroxylate a large range of hydroxyvitamin D derivatives available in my
laboratory. Some steroids which are structurally similar to vitamin D3 will also be tested. HPLC will be used to
detect product formation and if products are detected reactions will be scaled up to permit sufficient product to
be made to enable their identification by mass spectrometry and NMR (to be carried out by collaborators).
Subsequent studies will involving testing the biological activity of products.
2. Expression and characterization of the 20-hydroxylase, CYP314A1
CYP314A1 is a mitochondrial P450 that catalyses the 20-hydroxylation of ecdysone producing 20-
hydroxyecdysone, also known as insect moulting hormone. It is encoded by the gene known as shade, a
member of the Halloween family. This steroid hormone controls moulting of immature insects and
differentiation into pupae and adult. Thus CYP314A1 is a potential target enzyme for specific inhibitors to
control insect pests. CYP314A1 has some properties very similar to P450scc including catalysing 20-
hydroxylation of sterols, but the enzyme has not been purified for full characterization. The aim of this project
is to express CYP314A1 in E. coli, purify the expressed enzyme and study its ability to 20-hydroxylate
ecdysone. A range of other potential substrates including cholesterol and vitamin D will also be tested since their
20-hydroxy products have anti-cancer properties. HPLC will be used to measure product formation.
54
DR DANIELA ULGIATI Room 3.03, Bayliss Building, Phone: 6488 4423
Email: H [email protected] UH
My research interest is in the role of complement in health and disease. My ambition is to clarify the roles of
complement and B cell biology in autoimmune disease, using Systemic Lupus Erythematosus (SLE) as a model
for this and other autoimmune diseases. Specifically, my research focuses on the control of complement
receptor in health and disease. Students with a background in Molecular Biology, Biochemistry, Genetics or
Immunology are able to apply. Students will be exposed to a range of techniques including Genotyping,
Chomatin Assays, ChIP assays, DNA sequencing and cloning, cell culture, stable and transient transfection
assays, PCR, DNA binding assays, proteomic analysis, and FACS analysis.
PROJECTS
1. Isolation of Transcription Factors Involved in Regulating Human Complement Receptor 2
(CR2/CD21) during B Cell Development.
Complement receptor 2 (CR2) plays an important role in the generation of normal B cell immune responses as
demonstrated by CR2 knockout mice. As modest changes in levels of CR2 expression appear to effect B cell
responses, understanding the transcriptional control of CR2 is critical. More recently, a role for this receptor has
been established in the differentiation of normal B cells. Premature expression of CR2 resulted in marked
reduction in peripheral B cell numbers as well as mature B cells that are defective in their antibody responses.
This project involves the study of this gene during the B cell development process. Our analysis of the
transcriptional control of human CR2 show that this gene is complexly regulated by the presence of both
promoter and intronic silencer elements. Within these elements we have identified two regions critical for
transcriptional regulation. The first is a CBF1 binding site within the intronic silencer and the second is a cell
type specific repressor within the CR2 proximal promoter which binds E2A proteins as well as CBF1. Together
with these known transcription factors, many as yet unidentified proteins bind the functionally relevant sites.
This project involves studying the role of the identified factors during B cell development in vivo using
chromatin immunoprecipitation assays (ChIPs) and B cells lines that represent different stages of B cell
development. Isolation of and identification of the unidentified binding factors will be achieved using 2D
gel/proteomics based approaches.
2. The role of CR2 promoter polymorphisms in Systemic Lupus Erythematosus (SLE) and Rheumatoid
Arthritis (RA).
Complement receptor 2 (CR2) is an important receptor that is required for a normal B cell immune response. It
is expressed at a critical stage in B cell development and has been implicated in a number of autoimmune
diseases. The significance of mechanisms that regulate CR2 expression is apparent by studies of human B cell
CR2 expression in patients with SLE and RA. Both patient groups have abnormalities in the expression of CR2
on B cells (~50% of normal) and this decrease correlates with disease activity. With the recent advent of
transgenic and knockout mice, several groups have examined the importance of CR2 in a lupus prone mouse
model. Studies of these mice have also found an early decrease in CR2 expression that is initially detected prior
to any major clinical manifestations. We have recently sequenced the CR2 promoter in a number of SLE
patients and have found several single nucleotide polymorphisms (SNPs) within functional regions of the
promoter. We are currently assessing the functional implications of these polymorphisms on the transcriptional
regulation of CR2. This project involves determining the expression status of CR2 on patient B cells by
correlating cell surface expression with mRNA levels and transcriptional activity. Furthermore, collating the
expression and transcriptional data with the promoter phenotypes will ultimately determine whether these
promoter polymorphisms are indeed having an effect on CR2 expression in patients with autoimmune diseases.
55
3. Understanding the Role of Notch Signalling and associated Transcription Factors in Lineage
Commitment.
Notch signaling is an evolutionarily ancient mechanism which plays a critical role in dictating cellular fates.
Signals transmitted via Notch receptors control how cells respond to developmental cues and in turn control
lineage commitment. Notch signalling is intimately involved in lineage specification and differentiation of
lymphocytes.
Commitment to the B-lineage requires inhibition of Notch signals in lymphoid progenitors. Notch signals in this
context repress Pax5 expression thereby blocking B-cell differentiation. On the other hand, negative regulation
of Notch signals by the inhibitory Notch modulator deltex1, skews commitment of lymphoid progenitors to the
B-lineage. While, Notch1 signaling must be down-regulated to permit B-cell commitment, the involvement of
Notch signaling at subsequent stages of B-cell development in bone marrow have not been clearly defined.
Notch signaling also has important consequences for T lymphocytes. Dysregulated Notch1 signaling leads to T
cell leukemia in humans and mice. The ability of Notch to cause
T cell neoplasia results from aberent expression
during thymocyte development, where Notch receptor expression and signaling occur at distinct developmental
stages. There is evidence that Notch expression at very early stages
of lymphoid development commit
progenitors to the T cell lineage. Recent evidence indicates that Notch may also influence mature T cell
development.
We have recently developed an ex vivo model in which to study Notch signaling. Cells are co-cultured with
stromal cell lines ectopically expressing the Notch ligand, delta-like-1 (OP9-DL). Cells attached to the stroma
or in suspension following co-culture were harvested and can be analysed for differentiation and neoplastic
markers and associated transcription factors. Since Notch signaling is known to upregulate the bHLH factor
HES-1, we can also measure transcript abundance of this marker of Notch activation to ensure proper induction
of Notch by dela-like-1 ligand in the co-cultures.
4. Characterisation of the Upstream Repressor Element in the Complement C4 Gene and its control by
Lupus-associated Factors. (Co-supervised with Prof Lawrie Abraham)
The fourth component of human complement (C4) is a serum protein involved in initiation of immune and
inflammatory reponses. Previously, we have analysed the transcriptional regulation of the C4 gene. To
determine the requirements for basal and regulated expression, we have analysed the promoter region of C4 in
reporter gene assays, using deletion and mutant reporter constructs and in EMSA analysis. We have mapped a
number of promoter elements that are responsible for basal and interferon-gamma regulated expression. We also
discovered a novel two-part regulatory element within the promoter which appears critical for C4 expression in
hepatic cells. The reporter gene analysis results indicated the presence of repressor elements between –468 and
–310 (which contain putative binding sites for GATA and Nkx2) that had the effect of decreasing promoter
activity by more than 90%. In addition, these distal element/s appeared to be acting in concert with a complex of
Sp1/3 and BKLF-binding GT box elements around –140. This interaction has the effect of masking the very
strong negative effects due to the distal region. The mechanism for this masking effect is currently unknown, but
our hypothesis is that interaction with the –140 region prevents interaction of the upstream element with the
proximal basal elements (see Figure). We hypothesised that there would be an extracellular signal that regulated
C4 expression via this repressor element. In searching for such an agent we found an activity in serum from
Luus nephritis NZW X NZB F1 mice that was able to repress C4 transcription via the two-part element in the
C4 promoter. This project will involve the further characterisation of the repressor elements and the transcription
factors that interact with them, and a subsequent investigation of the mechanism of repression. Also, the
identity of the Lupus-associated factor will be investigated following purification.
56
Structure of the dimeric L-amino acid
oxidase from the snake venom of Malayan
pit viper. The glycosylations are also
indicated.
WINTHROP PROFESSOR
ALICE VRIELINK 31BRoom 4.31, Bayliss Building, Phone: 6488 3162
32BEmail: [email protected]
Protein Structure by X-ray Crystallography The studies in my lab focus on crystallographic analysis of a variety of proteins with the aim of using structural
analysis to better understand their biology. The structural biology laboratory is well equipped with state of the
art robotic crystallization equipment, X-ray diffraction equipment and computational facilities for structure
solution and analysis. Expression and purification resources are available in the laboratory in order to obtain
sufficient quantities of protein for crystallographic studies. In addition we carry out kinetic and spectroscopic
analyses to establish the quality of protein and pursue biochemical and biophysical studies to better correlate
function with structure.
PROJECTS
1. Endotoxin Biosynthesis in Neisseria.
The Gram negative bacteria, Neisseria meningitidis, is the causative agent of meningitis and is responsible for
significant mortality throughout the world. A characteristic feature of these bacteria is the presence of
lipooligosaccharide (LOS) molecules on their outer membranes. These complex molecules, also called
endotoxins, are structural components that play a role in bacterial immune evasion mechanisms hence present
interesting opportunities for the development of vaccines against the organism. A large number of enzymes are
involved in LOS biosynthesis including the additions of carbohydrate moieties to the endotoxin molecule and
enzymes involved in modification of LOS to provide it with the molecular features that facilitate recognition by
the host organism. A greater knowledge of the biosynthesis and regulation of meningococcal
lipoooligosaccharides will provide a more detailed understanding of the role of this molecule in pathogenesis
and disease. In collaboration with Professor Charlene Kahler of the Department of Microbiology and Dr. Keith
Stubbs of the Department of Chemistry at UWA we have begun a study to establish the structural and functional
relationships of these enzymes. Towards this aim, overexpression systems for these enzymes must be developed
in order to produce sufficient amounts of protein for structural and kinetic studies.
This project will involve cloning, protein expression, purification, crystallization and structure determination
using crystallographic techniques. Kinetic assays for the enzyme will be established in collaboration with Dr.
Stubbs and biophysical methods will be undertaken to characterize the protein. This project will be correlated
with functional studies carried out by Dr. Kahler and coworkers.
2. Studies of Snake Venom L-amino acid oxidase
L-amino acid oxidase is a flavoenzyme catalyzing the
stereospecific oxidative deamination of L-amino acids to give
the corresponding -keto acids. It is found in high
concentrations in a number of different snake venoms,
constituting up to 30% of the total venom proteins and is thought
to contribute to the toxicity of the venom. The enzyme has also
been shown to possess antibacterial, anti-HIV and antineoplastic
or apoptosis-inducing activity. The general mechanism of
cytotoxicity by the enzyme is thought to be due to the generation
of H2O2. Indeed, studies have shown that the addition of
catalase, a scavenger of H2O2, protects the cell from the toxic
effects of the enzyme. However other factors may also
contribute to the apoptotic activity including the glycosylation
moiety of the enzyme and an increase in the presence of
substrate. The structure of the enzyme in the presence of a
substrate and an inhibitor have been determined in our
laboratory and reveal a channel that may act as the peroxide exit
route from the active site. The channel exits near to the location
of one of the two glycosylation sites on the protein surface.
Further characterization of this enzyme and its mechanism of apoptosis will require production of wild type
enzyme as well as specific mutants, which affect catalytic activity. The protein is not able to be expressed in a
functional form in a bacterial expression system due to the presence of extensive glycosylation. Thus it must be
57
expressed in a eukaryotic system. Towards this aim a yeast expression system for the enzyme has been
established and provides a basis for production of both wild type and mutant forms of the protein for further
biological studies.
In this project you will use the yeast expression system to produce functional protein. Site directed mutagenesis,
kinetic analysis, crystallographic studies and apoptosis studies will be undertaken to establish the roles of
discrete residues in oxidation chemistry and its relationship to apoptosis.
3. Structural Studies of an Engineered Cephalosporin Acylase.
Cephalosporin C was originally isolated from the microorganism
Cephalosporium sp in 1945 as the first -lactam fused to a six-membered
ring. Thereafter a number of semi-synthetic analogues were developed
from the initial lead compound with fourth generation cephalosporins
being used currently. Many of the semi-synthetic analogues of
cephalosporin C (CephC) are synthesized starting with the conversion of
CephC to 7-aminocephalosporanic acid (7-ACA). This conversion
however involves a series of expensive chemical steps that require highly
reactive chemicals resulting in chemical wastes, which must be safely
disposed of. Hence altering the production method of semi-synthetic
cephalosporins to overcome these disadvantages is of great interest to the
pharmaceutical industry. An enzymatic method to produce semi-synthetic
cephalosporins from 7-ACA using D-amino acid oxidase and glutaryl-7-
amino cephalosporanic acid acylase is also possible and, although it
eliminates the problems associated with toxic waste products, it is
expensive and inefficient for industrial production. Therefore a one-step
conversion of CephC to 7-ACA is highly desirable. For this conversion,
utilization of glutaryl-7-amino cephalosporanic acid acylase (gl-7-ACA acylase) and altering its substrate
specificity and activity for the substrate CephC rather than glutaryl-7-amino cephalosporanic acid (gl-7-ACA)
offers an ideal solution. Towards these aims we are working with Prof Pollegioni (Universita degli Studi
dell‟Insubria, Italy) to design and characterize mutants of gl-7-ACA acylase with switched substrate specificity.
A double mutant of gl-7-ACA acylase (H296S-H309S) which exhibits 22 fold enhanced specificity and
reactivity of CephC over the natural substrate gl-7-ACA has already been designed. Our laboratory has
determined the crystal structure of this mutant and the wild type enzyme in order to establish the structural
consequences of the mutation that facilitate altered specificity.
The project involves further engineering of the active site through a mutagenesis approach to identify other
mutations that could enhance substrate specificity. The designed mutants will be prepared by site directed
mutagenesis, protein expressed, purified and crystallized. Substrate complexes of crystals will be prepared and
structures determined by X-ray crystallography methods.
Crystal structure of the H296S-
H309S double mutant of gl-7-
ACA acylase.
58
WINTHROP PROFESSOR
R JOHN WATLING Forensic Chemistry
Forensic Science Building, Phone: 6488 4488
Email: [email protected]
Forensic Chemistry Research Group Expertise and Interests:
The Group has two main research initiatives, firstly, spectral fingerprinting of crime scene evidence and
provenancing metals, projectiles, gemstones, glass, oriental ceramics, paintings, foodstuffs, explosives, plastics,
drugs and environmental materials, and secondly nano-forensics, a completely new area of forensic science
associated with the development of nano-sensors for real-time crime scene and terrorist activity investigations
by determining the presence of explosive gases, biological agents and residues.
Group Activities:
It is impossible to discuss in detail the diversity of projects being undertaken by the Forensic Chemistry
Research Group at UWA, however, any student wishing to obtain information should contact John Watling for a
CD of the group‟s activities.
Introduction:
With the increase in both sophistication and frequency of crime and the continuous decrease in Governmental
funding of police and law enforcement authorities it has become necessary for forensic chemists to be aware of,
to develop and to apply, relevant new analytical technology to assist them in "fast tracking" forensic
investigations. Furthermore, as criminals become more careful about leaving "debris" at a crime scene the
amount of evidentiary material is becoming smaller and
increasingly more difficult to analyze using conventional
analytical methodology. A significant setback for criminals
occurred with the advent of ICP-MS. This technique provides
an improvement in detection limits for most elements in the
Periodic Table of often more than three orders of magnitude
over conventional absorption and emission techniques.
Consequently it has now become more possible to obtain
analytical information for a wide range of elements on much
smaller samples. Incorporation of laser ablation with ICP-MS
has the potential to solve many of the existing problems
associated with provenance establishment of scene of crime
evidence as even the initial Nd-YAG lasers were capable of volatilization of relatively small craters (<100 m in
diameter) thereby removing often only a relatively tiny amount of the evidentiary material. The recent advent of
UV and Excimer lasers decreased the sampling volume to crater sizes of <10 m and thereby decreased the size
of potentially analyzable debris. The current research group in the application of lasers to forensic investigations
in a world leader in this technology and is a founder member of the international NITECRIME Network of
forensic mass spectrometric CSI laboratories.
59
The science of “Spectral Fingerprinting” is on its infancy and although recorded in case law in five countries
researchers have only scratched the surface of the technology. Consequently application of this technology is
suited to Honours, masters and PhD projects as well as considerable post doctoral research initiatives.
Therefore, while some overview project types are discussed in this document, rather than identify specific
projects in detail to students, the student is encouraged to use their imagination to identify areas where the
application of this technology is relevant and to suggest these to members
of the Forensic Chemistry Group. In this way it will be possible to tailor
specific projects of particular relevance to the student to suit student
interest and commitment. Suggestions such as the spectral fingerprinting
of Tapes and ties used in rape and drug transport, pencils and inks used in
forgeries, glass, pollen, plants, plastic rope, metals from crime scenes,
fibres, abrasive minerals, paper and canvass used in art forgery, statues,
clays, guns and projectiles are all relevant for consideration. Give it a
thought yourselves and come and see us. Current Honours students are
investigation the provenance establishment of diamonds, gold and
identifying the provenance of oil at ram raids and hit and run events.
PROJECTS
Some Possible Suggestions for Projects in Environmental Forensics:
1. The recent recognition of a lead problem in Esperance has resulted in an increase in interest in the
distribution of lead in the environment. Of particular risk are young children and babies. We propose to develop
a method of teeth analysis (lead is sequestered in teeth) to plot the history of lead intoxication by humans and to
look at methods of determining changes in the lead pollution of the environment with time. In addition we will
look at an Ibex tooth from the last European Ice Age ad determine if we can see the reflection of pasture
changes from summer to winter and tell how old the animal was when it died some 20,000 years ago.
2. The international requirement to provenance foodstuffs has led to the Forensic Chemistry Group at UWA
pioneering the inception of PROOF (The Australian and New Zealand Proof of Origin of Foodstuffs)
programme. This programme interfaces with the European equivalent programme (TRACE). We have projects
on developing methodology for the elemental fingerprinting of Milk Powder, Mineral Waters and Wine. We
even have some research dollars to buy some of the necessary ingredients! These projects will lay the
foundation of our involvement with the European programmes in these products and will compliment our
existing projects for tea and drugs.
Please remember that these are not the only projects on offer, they only from a basis for discussion towards a
relevant equivalent which can be mutually developed.
60
W/PROFESSOR JIM WHELAN ARC Centre of Excellence in Plant Energy Biology
Room 4.73, Bayliss Building, Phone: 6488 1749
Email: [email protected]
Molecular Genetics and Genomics
We use a variety of post-genomic approaches to carry out discovery based investigations concerning
the development and stress tolerance of plant model organisms, primarily rice and Arabidopsis. The main
projects running in the laboratory focus on the biogenesis and function of plant mitochondria, and on the role of
signaling events involved in plant phosphate uptake. Both mitochondria and phosphate metabolism are key
players in energy production in plants, making our investigations highly relevant for fundamental and applied
research. The students will be trained in a wide variety of molecular and cellular biology techniques, ranging
from gene expression analysis, quantitative proteomics and metabolomics, physiology to bioinformatic analyses.
This research is carried out in the ARC Centre of Excellence in Plant Energy Biology providing
students with training and hands-on use of state-of-the-art equipment. Previous students have received
international fellowships (EMBO, Human Frontiers, Australian Research Council) to carry out their own
research projects in Europe, USA and Australia. National and international research agreements with the
Australian National University, the University of Sydney, Zhejiang University (Hangzhou, China), The Max-
Planck Institute of Molecular Plant Physiology (Potsdam, Germany), Ludwig-Maximilian University (Munich,
Germany), and Umeå and Stockholm Universities (Sweden) provide students with the opportunity to study
overseas, supported by a variety of grants or scholarships provided by the Centre.
PROJECTS
1. Mitochondrial biogenesis and regulation
Mitochondria are key organelles in eukaryotic cells that play essential roles in energy production, various
biosynthetic pathways and in cell death. Thus mitochondria play key roles in the life and death of cells. As most
mitochondrial proteins are encoded in the nuclear genome and need to be imported into the mitochondria, the
biogenesis of mitochondria is a complex and well regulated process. The aim of our research is to characterise
the protein complexes that allow import of proteins from the cytosol to the mitochondria, the function of the
proteins involved in energy metabolism itself, and how these processes are regulated during development and
stress conditions. Using transcriptomic and proteomic approaches we have identified several novel
mitochondrial proteins that are important for mitochondrial function, but their mode of action is currently poorly
understood.
Mitochondrial dysfunction caused by a variety of environmental changes and stresses results in altered nuclear
gene expression in plants, called mitochondrial retrograde regulation. Overall this altered nuclear gene
expression results in mitochondria-mediated ability of the plant to cope with those stresses. However the
identity of the genes involved in mitochondrial retrograde regulation is largely unknown. We use genetic
approaches combined with state-of-the-art sequencing technologies to discover new genes that regulate the
ability of the plant to respond to stress. The overall aim is to analyse the functions of these different proteins and
pathways, thereby contributing to an integrated understanding of the biogenesis and regulation of mitochondria.
A variety of projects are available in this area. Each project has a post-doctoral team leader and 1 to 2
Ph.D students. Honours students working on these projects will join these small teams with their own individual
projects:
- Exploring the role of WRKY transcription factors in the molecular regulation of plant stress responses.
Dr Olivier Van Aken – [email protected]
- Studying the gene network that regulates mitochondrial stress response in Arabidopsis thaliana. Dr. Aneta
Ivanova - [email protected]
- Determining the functions of novel protein transporters. Dr Monika Murcha
([email protected]) and Yan Wang -([email protected])
- Investigating the role of temperature and climate zones in grape berry development using next generation
transcriptomic profiling technologies. Dr Estelle Giraud - [email protected]
- From seed to plant: understanding rice development on a transcript level. Dr Reena Narsai –
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- The mitochondrial bacterium 'The elephant in the room' - Exploring the adaptation of bacterial signalling
pathways for the successful development of higher plants. Owen Duncan - [email protected]
2. Using –omics and biotechnology to develop crops with improved phosphate use efficiency
Phosphate is an essential element for life, required for all energy-dependent processes in a cell, and as a
component of organic molecules, such as ATP, nucleic acids, membrane lipids and proteins. Plants have
developed various strategies to cope with the limited bioavailability of this element and the molecular
mechanisms that drive these processes have only recently started to become unravelled. Phosphate supply is of
particular importance for crops such as rice that are grown in nutrient-poor weathered soils. Current farming
practice requires application of large amounts of fertiliser derived from non-renewable phosphate rock. These
fossilised phosphate deposits are predicted to become exhausted in the next century and drive plant production
costs up, an issue of international economic importance as global demand for food increases and the
environmental damage of fertiliser application becomes apparent.
We use high-throughput molecular approaches to search for key regulators of the response to phosphate stress in
rice and the model plant Arabidopsis. Understanding the way plants adapt to a limiting nutrient environment
will allow us to develop novel biotechnology based solutions in cereal crops that can use the phosphate pool in
soil more efficiently and reduce the need for fertiliser application.
These projects are part of an international collaboration funded by the Australian Research Council
SuperScience program and the Chinese National Science Foundation between UWA and Zhejiang University.
Projects will be supervised by SuperScience fellows with expertise in –omic techniques, including
transcriptomics, proteomics and metabolomics.
- Using laser capture microdissection and next generation sequencing to analyse cell-specific responses to
phosphate starvation in rice. Dr David Secco – [email protected]
- Tracking phosphate stress-induced proteomic changes in rice with mass spectrometry. Dr Ralitza
Alexova – [email protected]
- Getting the message across: coordinating retrograde and anterograde signalling of mitochondrial protein
import upon phosphate deficiency in rice and Arabidopsis. Dr Marna van der Merwe –
References
Refer to http://www.plantenergy.uwa.edu.au/ for all publications and more details about scholarships.
Giraud E, Ng S, Carrie C, Duncan O, Low J, Lee CP, Van Aken O, Millar AH, Murcha M and
Whelan J (2011) TCP transcription factors link the regulation of genes encoding mitochondrial proteins
with the circadian clock in Arabidopsis thaliana. The Plant Cell 22:3921-3934.
Giraud E, Van Aken O, Ho L and Whelan J (2009) The transcription factor ABI4 is a regulator of
mitochondrial retrograde expression of Alternative oxidase 1a Plant Physiol 50: 1286-1296
Zheng L, Huang F, Narsai R, Wu J, Giraud E, He F, Cheng L, Wang F, Wu P, Whelan J, Shou H
(2009) Physiological and transcriptome analysis of iron and phosphorus interaction in rice seedlings
Plant Physiol 151:262-274
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ASSISTANT PROFESSOR
DUNCAN A. WILD Room 3.31, Bayliss building, Phone: 6488 3178,
Email: [email protected]
Laser Spectroscopy & Computational Chemistry
Research interests include: Spectroscopic investigations of gas phase ionic clusters, ab initio calculations to
predict infrared and photoelectron spectra, apparatus design and development.
5 projects are offered for prospective students. Projects 1-3 are concerned with spectroscopy of fundamental, yet
important, gas phase species using the TOF-PES apparatus. Project 4, in collaboration with Assoc. Prof. Scott
Stewart, deals with the synthesis and spectroscopy of novel carotenoids. Project 5 is theoretical in nature, and
involves modelling photoelectron and infrared spectra via ab initio methodologies.
PROJECTS
1. Photoelectron spectroscopy of atmospherically and astronomically important species
The spectroscopy projects are based on a time of flight (TOF)
mass spectrometer coupled to a PhotoElectron Spectrometer
(PES) which is now operational and churning out results in the
Wild Lab. The idea behind the experiment is:
1) Create exotic gas phase anion-molecule clusters.
2) Mass select a specific cluster using TOF mass spectrometry
3) Record a photoelectron spectrum using the fourth harmonic
of a pulsed Nd:YAG LASER ( = 266nm).
The rate and direction of chemical reactions is determined by the potential
energy surface governing the interactions between the species. Using
photoelectron spectroscopy of anion-molecule 1:1 complexes allows us to
probe the neutral potential energy surface. Spectra are shown to the left for
the chloride and bromide-carbon monoxide complexes.[1]
In this project, you will extend our studies to look at fundamental species
with Nitrogen and Sulphur containing molecules attached to an anion.
These species have relevance for the chemistry occurring in our
atmosphere, and that of distant celestial bodies. This project is flexible in
that you can choose the systems to investigate! We are currently
developing an oven source which will allow for more flexibility in our ion
production techniques.
2. The inception of solvation
Ever wondered what is occurring on the microscopic scale
when solutes dissolve in a solvent? What are the dominant
forces at play? How many solvent molecules are in close
contact with the solute, or in subsequent solvation shells?
Using the tof-pes we are in a position to answer these
questions!
With mass spectrometry we can probe one cluster size at a
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time and build a picture of size dependent properties, eventually seeing the closing of a solvation shell. In this
project you will record the photoelectron spectra of clusters of the form X-…(M)n with n=1,2,3,… and
supplement the spectra with ab initio calculations. We will target ligands such as C2H2 and CO as prototypical
solvent molecules. Shown in the figure are photoelectron spectra recorded recently of the I-…(CO)n with n=0-4.
The shift in the peak positions is determined by the intermolecular interactions between the solvent molecules
(CO) and solute (I-).
3. Electronic Spectroscopy of cation-molecule complexes Space is not empty!! In fact there are many regions which are dense with interesting molecules that so far have
not been unambiguously identified. Some potential candidates are polycyclic aromatic hydrocarbons, i.e.
naphthalene, anthracene, and so on. In this project you will create clusters between these molecules and Argon
cations, and then obliterate them with UV radiation.
The project will be run on the TOF-PES, however by operating it in cation mode rather than anion mode. The
photoelectron spectrometer will not be used, instead we will infer absorption of a photon by the neutral
dissociation products that result. This project will utilise our newly acquired laser system, which is a dye laser
pumped by a Nd:YAG laser, with a tunable range of 210-710nm (cool toys to play with!).
4. Synthesis and ultra-fast spectroscopy of novel carotenoids (co-supervised by Assoc. Prof. Scott
Stewart)
If you can‟t decide between a synthetic or physical chemistry project, then why not have the best of both
worlds? In this project you will be involved with the synthesis of novel apo-carotenoids, with nitrogen
containing functional groups. As part of the project you will collaborate with researchers at the University of
Sydney and utilise their femto-second laser system to record transient absorption spectra to determine the energy
relaxation pathways of these important molecules.
Carotenoids are prevalent in nature, and notably are found in the photosynthetic system. Their role is to both
protect the system from oxidative attack by singlet O2 and also to funnel energy into the PS system to aid
photosynthesis. Carotenoids feature alternating C-C and C=C bonds along a carbon back bone, with various
functional groups and ring systems attached at the ends.
5. Modeling photoelectron and infrared spectra of small dimer (1:1) complexes
Ab initio methods (calculations from first principles, i.e. no experimental
input) are used routinely to predict structures and energetics of molecules
and clusters (for some examples see reference [2] and citations within).
In this project you will model photoelectron of small dimer clusters. We
will start with basic approximations, and then extend to producing multi-
dimensional potential energy surfaces! (sounds impressive, heh?)
We have a healthy allocation of computing time with IVEC [3] and the
NCI [2] facilities. This project is ideal for those who are interested in
theoretical chemistry, spectroscopy, computing, code production, fooling
around with Unix(Linux), and working with multiple CPUs!
References:
1. K.M. Lapere, R.J. LaMacchia, L.H.Quak, A.J. McKinley, D.A. Wild, Chem. Phys. Lett., 504, 13-19 (2011)
2. D.A. Wild and T. Lenzer, Phys. Chem. Chem. Phys., 2005, 7, 3793-3804
3. http://www.ivec.org/ & http://nci.org.au/
Come by and see Duncan for more information, or drop by the lab to see “The Beast” (aka the TOF-PES)
and have a chat with students in the group: Kim Lapere (PhD), Marcus Kettner (PhD), and Stephen Dale
(Hons) about what life is like as a laser spectroscopist.
Potential describing the H-
bonded S-H stretching mode
of Cl- …H2S
64
PROFESSOR
MICHAEL J WISE 35B Room 2.09, Bayliss Building, Phone: 6488 4410
36B Email: [email protected] UH
Bioinformatics and Computational Biology
Research in the Bioinformatics and Computation Biology Lab. boils down to the application of computational
techniques to investigate biological questions. Current application domains include:
Bioinformatics of anhydrobiosis (species‟ ability to survive with minimal water)
Microbial bioinformatics
Low complexity/natively unfolded proteins
PROJECTS
1. Systems Approaches to Oxidative Stress (Jointly supervised with Assoc. Prof. Peter Arthur)
Oxidative stress is caused by reactive oxygen species (ROS) and is thought to exacerbate pathology associated
with many chronic diseases and conditions. Examples include Alzheimer‟s disease, atherosclerosis, dementia,
diabetes, emphysema, heart disease, HIV/AIDS, kidney disease, liver disease, muscular dystrophy, Parkinson's
disease, Rheumatoid arthritis, some cancers and aging. However, preventing the harmful effects of oxidative
stress is not a simple matter, as antioxidant treatments have generally been ineffective in the treatment of these
conditions.
One challenge has been the lack of understanding of the various molecular mechanisms by which oxidative
stress causes pathology. We have established that cysteine residues on proteins are particularly sensitive to
oxidative stress and our laboratory is now playing a leading role in identifying proteins sensitive to oxidative
stress. Our work, and the work of others, has established that multiple proteins are sensitive to oxidative stress,
which means oxidative stress could have a widespread impact on many cellular processes (metabolic pathways,
ion transport, protein synthesis, protein degradation, gene expression, signal transduction pathways).
The objective of this project is to develop and use bioinformatic methods to identify the cellular processes and
organelles that are particularly sensitive to oxidative stress. This will involve categorizing the involvement of
proteins (those identified as sensitive to oxidative stress) in different cellular processes. You will be using
pathway analysis software such as IPA (www.ingenuity.com), keyword clustering software (Protein Annotators
Assistant) and databases such as BioCyc, Reactome and Kegg to look for common themes/processes. Protein-
protein interaction data and data about predicted location may also be useful.
2. Viral Codons
You are no doubt aware that the "Universal" codon translation table in fact only applies to eukaryote genomes,
and even then not to all of them; slime mold has a different table. The set of different tables can be found at:
http://www.ncbi.nlm.nih.gov/Taxonomy/Utils/wprintgc.cgi?mode=c If you look at that site you will notice that
there is no mention of viruses. One may assume, however, that because viruses are dependent on the replication
machinery of their hosts that their genes will be encoded like their hosts, i.e. use the same codon translation
tables. So, for example, MUMPS will use the Universal table, while lambda phage will use a bacterial table.
The Codon Adaptation Index was developed some years ago and reflects the observation that some codons are
far more used than other codons for a given amino acid, arguably reflecting greater numbers of the
corresponding anti-codons. The authors also observed that highly expressed genes tend to use the most abundant
codons. The Codon Adaptation Index was developed to reflect these observations.
The project is to examine viral genes in terms of their Codon Adaptation Index to gauge the extent to which the
codon usage biases of a virus mirror that of its host. Is it possible to see significant differences between codon
usage in the different isolates of the same virus which target different species, e.g. influenza virus affecting
humans and birds.
65
3. Is Genome Plasticity a Cofactor of Microbial Virulence?
The Sit-and-Wait hypothesis of microbial pathogenicity for non-vector-borne pathogens (Walther and Ewald
2004) suggests a correlation between the durability of a non-vector borne microorganism and its pathogenicity.
(See also the review: Brown et al. (2006).) Under the hypothesis, durability – the ability to survive the stresses
associated with existing for a period outside a host – is, in effect, a cofactor for pathogenicity, in concert with
the necessary presence of conventionally understood virulence factors. That is, without an assortment of
virulence factors, a microorganism is unable to colonise a host, but if the microorganism is labile, virulence will
be tempered over time because an immobilised infective host is unable to move and thus unable to spread the
infection. In other words, durability genes – like vector based transmission – give the pathogen “other options”
beyond the survival of the host. An extension of this thesis is to include long-term dudrable energy storage as a
cofactor for pathogenicity because unless an energy store has been maintained the organism may have survived,
but it will not have the energy to produce the range of invasion mechanisms it requires, such as pili. In this
project you are to examine another possible cofactor: genome plasticity. Bacterial with plastic genomes leave
themselves open to being parasitized. On the other hand, having a plastic genome gives the organism other
options, in this case import of useful genes from other organisms, e.g. coresident in a biofilm. The overall aim of
the project is to find any protein coding genes that may enable greater plasticity, linking these firstly to the
difference organisms and then also to published mortality data as a proxy measure for virulence.
4. A Novel Method for Building Phylogenetic Trees
Phylogeny is the study of the relatedness of species. The way this is done these days is through the
computational analysis of genes in living organisms. The phylogeny of organisms is often depicted as
phylogenetic trees and there is a considerable literature on how best to create such trees. Most methods take as
input data from a single gene or protein sequence across a range of taxa. That is, the same gene is found in all
the species of interest and then compared to build the tree. The problem with this approach is that it assumes that
the gene is "typical" and that evolutionary pressures have acted in the same way across all the species to shape
that gene. A second problem is to find a gene that is both ubiquitous and conserved in its function, but with
sufficient variability to differentiate the various species possessing that gene. In this project you will create an
application which takes as its input the models generated by an existing genome analysis application as it
traverses whole bacterial chromosomes. Then, after normalising the elements of the data vectors, you will try
different methods for building phylogenetic trees from the data. In other words, rather than trying to find the
ideal gene around which to build a tree, this method will compare summaries of all the data available in
chromosomes or, by extension, entire genomes.
5. Low Complexity Protein Domains in Bacteria
Globular proteins, e.g. enzymes, have sequences whose sequences appear to be random. That is, at any point in
the sequence it is hard to predict what the next amino acids will be based on those you have seen to this point.
These are called high complexity sequences. Low complexity proteins and protein domains, on the other hand,
are peptide sequences whose compositions appear to be far from being random. A well-known example is the
tandem GPP repeats found in collagen sequences. Amino acid stutters (tandem repeats of the same amino acid)
are another type of low complexity sequence. In eukaryotes, low complexity proteins are often found in
structural proteins, such as collagen and mucin in vertebrates and glutenins in plants. Low complexity proteins
are also associated with a number of diseases, e.g. Huntingdon‟s disease is due to a pathological expansion of a
poly-glutamine stutter. Low complexity proteins are also often natively unfolded – they have little or no
organised structure at ambient temperature and pH, but may nonetheless still be functional. A survey in Wise
(2002) found that low complexity sequences are rare in bacterial and phages, but more common in eukaryotes
and their viral parasites. However, low complexity bacterial proteins do exist in bacteria, so the task in this
project will be apply predictors of low complexity and natively unfolded proteins to a range of bacterial
proteomes to determine where such domains are found and are there any functions that are associated with
bacterial proteins which have low complexity or natively unfolded domains. Further more, to what extent do the
archael proteomes follow any trends you observe in bacterial proteomes.
66
PROFESSOR GEORGE YEOH Room 2.59, Bayliss building, Phone: 6488 2986
Email: [email protected]
Liver Research Group Our research group focuses on the biology of the liver progenitor cell (LPC) called an ―oval cell‖ which
describes its shape. We envisage an enormous potential for this cell as the vehicle for cell and gene therapy to
treat liver disease. We contend it is superior to other cell types such as the hepatocyte and the embryonic (ESC)
or adult stem cell (ASC) for many reasons. In particular, it is robust and simple to freeze and store, then thaw
and grow by in vitro culture when required. It can be differentiated into either hepatocytes or cholangiocytes
(bile duct cells) quite easily and rapidly when maintained under appropriate conditions, therefore it is more
versatile than the hepatocyte. Most importantly, the LPC is developmentally close to the hepatocyte and the
cholangiocyte in contrast to the ESC or ASC, which will require many more steps and much coaxing to produce
useful cells for liver therapy. Our long-term vision is to hasten the day when human LPCs are utilised to treat
liver disease, especially end-stage liver disease for which currently organ transplant is the only solution. A
realistic expectation in the short term is to use LPCs to ―bridge‖ patients thereby extending their survival and
enhances their probability of finding a suitable organ donor. A more ambitious and longer-term aim is to use
these cells to circumvent the requirement for organ transplant. This may be possible with some liver diseases.
To utilise LPCs we must identify and understand the action of growth factors and cytokines, which influence
them. To accomplish this, we have characterised the pattern of cytokine expression in two mouse model of liver
disease that induces the appearance of LPCs. These studies indicate that a subset of inflammatory cells, the
macrophages and cytokines they produce namely TNF alpha and TNF like weak inducer of apoptosis (TWEAK)
are LPC regulators. To understand both the cellular and molecular mechanism of action mediated by
inflammatory cells we are using cultures of LPCs and LPC lines. This knowledge can be used to increase their
contribution to liver regeneration in vivo which can lead to positive outcomes for liver disease patients. Both in
vivo and in vitro, extended growth of LPCs results in transformation to cancer; in this context hepatocellular
carcinoma. Therefore it is important to document changes in gene expression that are responsible for
transformation. Recent developments in our laboratory which underpin the projects on offer are:
1 Isolation and characterisation of LPCs from adult human liver
2 Establishment of LPCs from a transgenic mouse which expresses beta-galactosidase when it becomes a
hepatocyte and LPCs which express EGFP which facilitates cell tracing.
3 Acquisition of the Cellavista instrument which allows for progressive, accurate, high throughput and
comparative growth characteristic of multiple cell cultures
4 Identification of chromosomal alterations (See Fig 1) and gene expression pattern differences between
normal and transformed LPCs as a result of expression profiling.
Accordingly research projects will exploit these new developments for they are designed to increase our
understanding of LPCs and establish their utility for treating liver disease.
Fig 1: (A) Chromosomal alterations during culture of an LPC line (BMEL) at passage 5 (A), 10 (B) and 15 (C).
Chromosome loss (red arrows), gain (blue arrows) and consistent mars (small arrowhead) seen between
passage 5 and 10. The chromosome in passage 5 and 10 remain telocentric, consistent with normal mouse
structure. The cells are hypotetraploid, however, there are less than 4 copies of chromosomes 4 (red dotted
circle) and more of chromosome 9 (blue solid circle). Massive transformation of chromosome structure has
occurred between Passages 10 and 15 and typical mouse chromosomes can no longer be identified. The
chromosomes have been assembled into a karyotype using traditional cytogenetic methods. They are grouped
according to size and similar banding patterns then arranged from largest to smallest. Structural changes
include duplication/translocation and increase in mars.
67
GENERATING FUNCTIONAL LIVER CELLS FROM LPCs
Assessing the ability of LPCs to synthesise urea
Ornithine transcarbamylase (OTC) is a urea cycle enzyme that is mutated in individuals with a metabolic
disorder - OTC deficiency. The consequence of accumulating ammonia affects many tissues and the liver
particularly is damaged. The condition affects young children with neurologic consequences, hence liver organ
transplant is necessary to treat those severely affected. Cell therapy using normal hepatocytes may also be
possible, but hepatocytes are difficult to maintain and store; and once transplanted may not survive for very
long. In contrast LPCs are robust and have the added advantage of long-term survival and the ability to
proliferate and continue to generate hepatocytes in situ.
This project evaluates the effectiveness of utilising LPCs to treat OTC deficiency. First, the ability of LPCs to
express OTC following differentiation into hepatocytes will be determined. Then their ability to synthesise urea
will be compared with hepatocytes.
Availability of the Spf-ash mouse model of human OTC deficiency through our collaboration with Professor Ian
Alexander of the Childrens Medical Research Institute in Sydney allows us to directly test our LPC lines in
these mice. This will be undertaken if the cells induce OTC and acquire the ability to synthesise urea following
differentiation.
We are also attempting to generate LPC lines from the Spf-ash mouse. They will serve as negative controls for
the differentiation studies. The CMRI group will use these cells to establish methods to correct the gene
deficiency in OTC-/-
LPCs as proof of concept studies in advance of applying this method to children with OTC
deficiency.
WHAT MAKES LPC’s BECOME CANCEROUS?
Comparing tumorigenic and non-tumorigenic LPCs
LPC lines have been established from p53 -/- as well as +/+ mice. Some grow in soft agar and produce tumours
when injected subcutaneously into nude mice; some do not. We are defining the differences beween these cell
lines at the molecular and cellular level to identify features which are causative and those which are
consequential in terms of cancer. Specifically we are documenting chromosomal changes and focusing on
oncogene candidates raised by gene profiling. Two anti-apoptotic genes IAP and Yap are prime suspects and
their expression at the mRNA level (through qPCR) and protein level (by Western Blot) are being be defined for
a range of cell lines and during tumorigenesis during culture. Current studies follow changes in LPCs as they are
passaged and progressively become tumorigenic. We are also documenting changes in expression of p53 and the
level of its activity by measuring the expression of downstream genes such as p21. We are also testing the
effects of culture conditions on tumorigenesis. In particular, we will determine whether the level of oxygen and
the composition of the culture medium with respect to growth factors contribute to transformation.
Does the level of ROS contribute to transformation of LPCs as they are maintained in culture?
LPC lines which are initially non-tumorigenic will become tumorigenic following repeated passaging in culture.
This project tests the hypothesis that ROS is an important contributor to the mutagenic events by passaging cells
in 20% O2 and 2% O2. It predicts that cells maintained in hypoxic conditions will less readily transform.
Another approach is to maintain cells in the presence of antioxidants (vitamin C or desferrioxamine) which
should produce the same outcome. Alternatively, the hypothesis would be also be supported if it can be shown
that transformation will occur sooner if cultures are maintained under conditions which produce higher levels of
ROS such as in the presence of ethanol or H2O2.
The tumorigenic state of the LPCs will be assessed by their capacity to grow in soft agar. This can be confirmed
by their ability to form tumours in nude mice. TBARS assay will be used to ascertain the ROS levels under
different experimental conditions. Tumorigenic LPCs also display gross chromosomal abnormalities and this
can be documented by karyotyping the resulting cell lines.
68
HOW TO APPLY
UWA Applicants
If you completed your undergraduate studies at UWA you should lodge an on-line
application via StudentConnect by clicking on the Apply for Honours link in the left hand
menu bar of StudentConnect.
Applications will open online on Monday 10th
October and close on Tuesday 20th
December.
Non-UWA Applicants
If you have not previously been enrolled at UWA, you apply through one of the following
centres, depending on your circumstances.
Domestic Students
Australian citizens, permanent residents and/or holders of a humanitarian visa or New
Zealand citizens apply through the UWA Admissions Centre via the UWA‟s Online
Application System (OASys).
International Students
International Students apply through the UWA International Centre.
Honours Project Preference Form
All applicants must complete the BBCS Honours Project Preference Form and return it to the
MCS Building reception by Friday November 11th
2011.
69
Biochemistry & Chemistry
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PPRROOJJEECCTT PPRREEFFEERREENNCCEE FFOORRMM
The purpose of this form is to ascertain your interest in our Honours/GradDipSci courses. It is appreciated that students may be exploring Honours/GradDipSci in more than one discipline. Phone the BBCS School Office (6488 4402) to be referred to the appropriate Coordinator to discuss any questions you may have.
PPlleeaassee rreettuurrnn ffoorrmm ttoo BBBBCCSS SScchhooooll OOffffiiccee bbyy FFrrii 1111tthh NNoovv 22001111
I am interested in Honours/GradDipSci in 2011 within the Discipline of:
Biochemistry & Molecular Biology Biomedical Science
Chemistry Genetics Nanotechnology
Note: You need to fill out a separate form for each Discipline if you are considering projects in more than one. Include projects for any Programme (e.g. Genetics, Chemistry, Biomedical Science etc) that will be located within one of the above Disciplines
I am considering mid-year entry to Honours in 2012
I am considering deferring Honours until 2013
I will will not be available for interview during the week 5 December - 9 December 2011
1. CONTACT DETAILS
Name…………………………………………………………………………………………………………………………
Address(es) (during period November/December 2011 – January 2012): ……………………………………………………………………………………………………………………………...…
………………………………………………………………………………………………………………………………...
Phone No (during same period) ……………………………..…………………………………….………...
Mobile No (during same period) ……………………………………………………….……………………..
Email address …………………………….………………………………………………..
2. PROJECT PREFERENCES
In order of preference:
1 Project No [ ] Supervisors …………………………………………………………… 2 Project No [ ] Supervisors …………………………………………………………… 3 Project No [ ] Supervisors …………………………………………………………… 4 Project No [ ] Supervisors …………………………………………………………… 5 Project No [ ] Supervisors …………………………………………………………... 6 Project No [ ] Supervisors …………………………………………………………...
If there are any points you would like us to take into consideration please note them below: ………………………………………………………………………………………………………………………………………...……………………………………………………………………………………………………………………… Signature………………………………………………………Date………………………………………………………
The Faculty’s End-on Honours on-line application form must be completed by December 20th 2011. Prospective candidates will be interviewed 5 December - 9 December 2011, although other arrangements can be made if candidates are unavailable. Those students who have submitted this project preference form and who are eligible to enrol in the course will be emailed a confirmation of eligibility as soon as exam results are known [approximately 20 December], and allocation of projects will be advised as soon as possible after this. Student Administration will send you an Authority to Enrol letter in January 2012.
Faculty of Life and Physical SciencesThe University of Western AustraliaM310, 35 Stirling HighwayCrawley WA 6009Tel: +61 8 6488 4402Fax: +61 8 6488 7330Email: [email protected]
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