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8/4/2019 The Operon
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BY:PRIYANKA PATEL
Msc.(B.T) SEM-I
ROLL NO:47
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Cells must onlyexpress genes when needed Geneexpression (transcription, translation)
takes uplargeamounts of cellularenergyandresources
Cells live frugallifestyles they conserveenergyandresources
So genes will only beexpressed when theirproducts are needed.
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Bacteria control genes at the transcriptional
level
In other words, the gene is either transcribed or
not, based on certain external stimuli
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Francois Jacob & Jacques Monod
first to describe operon system
coined thephrase operon
2005-2006
1961 /1965
Francois JacobJacques Monod
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An operon is a collection ofprokaryotic genes
transcribed together on a single mRNAtranscript to servea singlepurpose
Composed of
An operator, an on-off switch
A promoter
Genes for metabolic enzymes
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A Region of DNA that interects with a
repressorprotein to control theexpression of
a gene ora group of gene Found within thepromoterregion or
between thepromoterand geneencodingregions.
On-off switch to gene
Controls theaccess of RNA polymerase +
Gene
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A DNA sequenceat which RNA polymeres
may bind ,leading to initiation of
transcription Near the operator
Onepromoter controls transcription ofallgene in operon
Transcribedas 1 unit & a single mRNA ismade
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It code for theenzymes themselves. The structural genes ofan operon usuallylies
adjacent to oneanother,and the RNA
polymarase moves from one structural gene tothe next , transcribing all of the genes into asingal mRNA.
This extended mRNA is then translated into the
various individualenzymes of the metabolicpathway. Turning on one gene turns on all theproducing
gene ofan operon.
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Catabolic operon:An operon composed of
genes whoseproducts degrade organic
compounds.
Biosynthetic operon:An operon composed ofgenes whoseproducts are involved in
synthesizing compounds,such as amino acidsor vitamins,rather than degrading them.
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Inducer: A signal molecule that, when bind to
aregulatoryprotein, produces an increase in
theexpression ofa given gene
Repressor: Theprotein that binds to theregulatory sequence or operator fora
gene,blocking its transcription
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Corepressor:The small molecule that
associate with an aporepressor to form an
activerepressor.(e.g.,tryptophan is thecorepressor of the trp operon).
Aporepressor:A repressor in an inactive form
,without its corepressor.
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A repressible operon is one that is usually on;
binding ofarepressor shuts off transcription
Thetrp operon is arepressible operon An inducible operon is one that is usually off;
a molecule calledan inducer inactivates therepressorand turns on transcription
The classic example ofan inducible operon isthelac operon
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Inducibleenzymes usually function in catabolic
pathways; their synthesis is induced bya
chemical signal
Repressibleenzymes usually function inanabolic pathways; their synthesis is repressed
by high levels of theendproduct
Regulation of thetrp andlac operons involvesnegative control of genes because operons areswitched off by theactive form of therepressor
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Some operons arealso subject to positivecontrol through a stimulatoryprotein, such as
cataboliteactivatorprotein (CAP), an activator
of transcription When glucose (apreferred food source ofE.
coli) is scarce,CAP is activated by binding with
cyclic AMP ActivatedCAP attaches to thepromoter of the
lacoperon and increases theaffinity of RNA
polymerase, thus accelerating transcription
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When glucoselevels increase,CAP detachesfrom thelac operon, and transcription returns
to a normalrate
CAP helps regulate other operons that encodeenzymes used in catabolic pathways
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Thetrp operon contains 5 structural genes
,TRP -E,D,C,A,B.
These geneencode for 3 enzymes that turnchorismic acid into Tryptophan.
The operon is not expressedwhen the cellcontains sufficient amounts of tryptophan.
The operon is expressed when levels oftryptophan arelow.
18
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Thetrp operon is negativelyregulated by thetrp repressor protein
trp repressor binds to the operator toblock transcription
binding ofrepressorto the operatorrequiresacorepressorwhich istryptophan
low levels of tryptophan prevent therepressor from binding to the operator
19
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DNA
Tryptophan (co-
repressor)
TrpR protein
subunits
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Converts chorismate to anthranilate usingammonia.
Anthranilate is an amino acid that is laterbroken down into another amino acid,Trysoine.
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A type ofGlycosyltransferase thatparticipates in the synthesis of Tryptophan
Produces an amino acid calledAnthranilatewhich ,by the help of TRP E, is broken downinto anotheramino acid, Trysoine.
Converts -(5-phospho-D-ribosyl)-
anthranilateanddiphosphate toanthranilateand 5-phospho-alpha-D-ribose1-diphosphate.
It is 1 of the 2 components that together
encodeAnthranilate synthetase
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Involved in the synthesis of tryptophan and
amino acids.
It is a type oflyase that cleaves carbon-carbon bond.
this abilityallows it to converts 1-(2-
carboxyphenylamino)-1-deoxy-D-ribulose5-phosphate to C(1)-(3-indolyl)-glycerol 3-
phosphateand carbon dioxideand water
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Thereare two beta subunits which form a
dimer
Betadimer catalyzes the formation of L-tryptophan from L-serineand 1-(indol-3-
yl)glycerol 3-phosphate
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Tryptophan synthase is an enzyme found in
plants and bacteria
Thereare two separatealpha subunits They catalyze the formation of indoleand
glyceraldehyde 3-phosphate fromindoleglycerolphosphate in tryptophan
biosynthesis, which is needed by the Bsubunits.
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Tryptophan present
Regulator Gene Promoter Operator Structural GenesAttenuator
trpRmRNA
RNA Polymerase
NOTRANSCRIPTION
TrpR protein
(homodimer)
+ tryptophan
(corepressor)
TrpR aporepressor
+ corepressor(can
bind to operator)
Q: Why might the cell want
to produce an aporepressor
that is only activated by the
operons end product?
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Tryptophan Absent
Regulator Gene Promoter Operator Structural GenesAttenuator
trpRmRNA
RNA Polymerase
TRANSCRIPTION
TrpR protein
(homodimer)TrpR aporepressor
(cannot bind to
operator)
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Attenuator:A region of DNA upstream from
one or more structural genes, wherepremature
transcription termination(attenuation)can occur.
The Attenuation Mechanism oftrp Operon
Mechanism of Attenuation
When Ribosomereaches Trp
codon
Trp is scarce
Trp is abundant
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Attenuation model in Trp starved cells.
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Attenuation model in Trp non-starved cells
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Generality of Attenuation
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TheTrpR repressor negativelyregulates not
only the transcription of the trp operon but
also the transcription of its own gene,trpR. In theabsence of theTrpR protein,the
transcription of the trpR gene is about 5times higher then in thepresence of TrpR.
When theproduct ofa generegulates the
expression of its own gene,it is called
autoregulation.
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L- ara operon was the first e.g. ofpositiveregulation in bacteria to bediscovered.Its
usually called theara operon. Theara operon contains 3 structuralgene,ara-A,B,D together with theI andOsites.
The gene of this operon areresponsible forconverting L- arabinose into D-xylulos-5-phosphate,which can be used by otherpathways.
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The operon is not expressedwhen the
arabinose is absent in the cell.
The operon is expressed when thearabinoseis present in the cell.
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P
t
Pt
araC araB araA araD
Activator/Repressor
I1 I2
AraCdimer
CRP
O2 O1
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araO1:Itis an operator site. AraC binds to this siteandrepresses its own transcription from thePCpromoter. In thepresence ofarabinose, however,AraC boundat this site helps to activateexpression
of thePBAD promoter. araO2:Itis also an operator site. AraC boundat this
site can simultaneously bind to thearaIsite torepress transcription from thePBAD promoter
araI:Itis also the inducer site. AraC boundat thissite can simultaneously bind to thearaO2 site torepress transcription from thePBAD promoter. Inthepresence ofarabinose, however, AraC boundat
this site helps to activateexpression of thePBADromoter.
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CRP binds to theCRP binding site. It does not
directlyassist RNA polymerase to bind to the
promoter in this case. Instead, in thepresence ofarabinose, it promotes the
rearrangement ofAraC when arabinose ispresent from a state in which it represses
transcription of thePBAD promoter to one inwhich it activates transcription of thePBAD
promoter.
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ara A :Encode L- Arabinose isomerase whichconverts L- Arabinose into L- Ribulose.
ara B :Encode L- Ribulosekinase whichphosphorylates L- Ribulose to L-Ribulose-5-
phosphate.
ara D :Encode L-Ribulose-5-phoshate
epimerase which converts L-Ribolose-5-phosphate into D-Xylulose-5-phosphate.which can then be metabolized via the
pentosephosphatepathway.
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When arabinose is absent, there is no need to
express the structural genes. AraC does thisby binding simultaneously to araIandaraO2.
As aresult the intervening DNA is looped.
These two events blockaccess to thePBADpromoter which is, in any case, a very weak
promoter (unlike thelacpromoter):
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Arabinose absentNegative control
Blocked
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When arabinose is present, it binds to AraC
andallosterically induces it to bind to araIinsteadaraO2. Ifglucose is also absent, then
thepresence ofCRP bound to its site
between araO1 andaraIhelps to break theDNA loopandalso helps AraC to bind to araI:
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Arabinose present Positive control
araB A D
O1 CRP araI
araC
PC PBAD
O2
Transcription
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AraC can bind to three sites (araO, araI1, andaraI2) with different affinities
46
Fig. 15.23
(a) No arabinose present:
When AraC is bound to araO and toaraI1, looping of DNA occurs and
prevents transcription
(b) Arabinose present:Arabinose causes allosteric change in
AraC so that it cannot bind to araO
AraC interacts with RNA polymerase
only when both araI1 and araI2are
occupied
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TheAraCprotein not onlyregulate theara operon
but also negativelyautoregulates its own
transcription. If the concentration of AraC becomes too high,its
synthesis willagain berepressed.
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Thereare several mutations that couldpreventtheexpression of thearabinose operon genesandeven cause the cell to die
. A mutation in the araA genewill cause thebacterial cell to becomearabinose negative. Thismeans that the bacterium can no longer usearabinoseas a carbon source. It will not utilize itif the cell is grown in a minimal media containingarabinose,.
A mutation in the ara B genewillalso result inthe same state. However, amutation in the araD genewillresult in cell death.
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After the ara B geneproduct degrades L-ribulose to L-ribulose-5-phosphate, epimerase isnot synthesizedand cannot breakdown L-
ribulose-5-phosphate which accumulates in thecell. L-ribulose-5-phosphateis toxic to the cell when
present in high levels. Mutations couldalso occurin the araCgene
causing thepromoters, PBAD and Pc, to becomeinactiveand thearabinose operon remainspermanently repressed.
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It is also regulated through cataboliterepression , sothe gene forarabinose utilization are not expressed ifthe medium contains a better carbon source.
CAP may help open theloop of DNA created whenAraC bind to araO2 andara1.opening theloop mayprevent AraC from binding to araO2 andaraI1,facilating the binding of AraC to araI1 andaraI2 andtheactivation of transciption from pBAD.
Thus, the nt of glucose or another carbon sourcebetter then arabinose enhances the transcription ofthe ara operon.
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Scientists modified thearabinose operon in
pGLO to express theGFP gene
TagCells (to detect specific cells) Act as areporter gene
Source ofbioluminescence whenexposed to
UV light
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Molecular Biology of theGene:Watson et al.
MolecularGenetics of Bacteria: Synder &
Champness www.google.in
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