Center for Genomic Experimentation and Computation
Orna Resnekov
Molecular Sciences Institute
Berkeley, CA
Magritte
Goal: To combine genomic and computational research in order to make predictive models of biological systems.
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a
a factor
factor
zygotea/
The mating pathway
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a
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zygotea/
The Alpha Project
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Pathway after pheromone
We are trying to both model and simulate the pathway
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Simplified view of pathway
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Abundance (Ste5 ~500, Fus3 ~20,000)Reaction ratesAffinities of proteins for one anotherStructural information (not covered in our
current Center grant)
To simulate we need numbers…
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Key questions
What is the oligomerization state(s) of purified active Ste5?
What is the stoichiometry of the different multiprotein complexes that have kinase activity?
Which of the structures are biologically relevant Ste5 complexes?
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Complexity of the sample and purification status
Ste5, Ste11, Ste7, Fus3We have expression constructs to make
individual proteins and reconstitute the pathway
We are also purifying complexes from cells using affinity tagged proteins (TAP)
We have polyclonal abs, we are identifying posttranslationalmodifications (MS), we are making monoclonal abs to modified proteins, we have fluorescently tagged proteins.
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Hurdles to overcome
Need a good partner for structural studiesWe would like to be doing both cryo-EM
and crystallographySufficient material for these studies
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Resources needed
RO1 level funding for 3-5 people
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Response to pheromone
Bright field image Fluorescent image
2 hour pheromone treatmentReporter: Prm1-YFP
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The Team:Kirsten BenjaminRoger Brent-DIRECTORIan BurbulisTina ChinAlejandro Colman-LernerAndrew Gordon (P)Larry Lok (Math)Joyce MaccabeaGustavo PesceDavid PincusOrna ResnekovDan RockwellEduard SerraDavid Soergel (CS)Kouichi TakahashiKumiko YamaguchiRichard Yu (Struct. Bio.)Myron Williams
PNNLRichard Smith (Chem)Robert Maxwell
SBMLAndrew Finney
U. WA.Dee Dee MeldrumMark Holl
MITDrew Endy (ex MSI)Ty Thomson (BioEng)
CaltechShuki Bruck (CS)Mark ReidelMatt Cook