**CHAPTER 11 Regulation of Gene Expression in Bacteria
One DNA-binding protein, AraC, serves as an activator and as a repressor The switch is thrown by arabinose.
(a) Activation CAP+cAMP bind so ↓ glucose so Arabinose Operon = turned on-can bind Activator+ RNA Polymerase AraC protein+Arabinose= effector will bind AraC protein will turn on genes to Utilize lac Operon (b) Repression AraC protein changes conformational shape when no Arabinose binding araO binds to 2nd site opened up So it= repressed Ara genes are repressed
Leader peptides of amino acid biosynthesis operons
(a) early on AA are encoded (c) front loading protein lets know how wmuch protein =there
Abundant tryptophan attenuates transcription of the trp operon Only in Bact. Attenuation (a) Stemloop 2ndary structure of RNA- Intramolecular base pairing can toggle b/w 2 forms
(b) ↑ Trp level mRNA= being made The 3-4 stemloop= Rho Independent will cause transcription to stop There is going to be a lot of charge tRNA inserting Trp, only a small amt Trp= made-not functional (c) ↓ Trp level mRNA= made no tRNA with Trp the alternative 2-3 stemloop is made= not a transcriptional termination signal. Transcription continues.
Sigma factors control clusters of unlinked genes Genes are not lined up How get all 3 genes turned on at the same time? 3 genes all involved in the same process in bact so have same seqence in -10 to -35 regions They all have the same sequence in their proteins Reg protein A binds to all the promoters at the same time b/c all promoters are the same Most genes cause chemotaxis-bact goes toward food away from repellents
1 protein A =made with many sigma factors, each = used at different times to regulate different genes. So all genes around chr can be regulated at the same time Ex Ja= A housekeeping gene=constitutive=always on Both Jo +Jc =turned on by Ja when it is hot Jo= Heatshock protein Jc= sporulation
2 main mechanisms for regulating transcription of operons functioning in Different amino acid biosynthesis
1.Global control of operon mRNA expression 2.Fine-tuned control
Cells need mechanisms that fulfill 2 criteria: They must be able to:
1.recognize environmental conditions in which they should activate or repress the transcription of the relevant genes 2.Toggle on or off, like a switch, transcription of each specific gene or group of genes Genetic switches
Dual Positive and Negative Control: The Arabinose Operon Map of the ara operon
AraC protein= Control gene
Bacteria can either: - Acquire nutrients they need from the environment - Synthesize them using enzymatic pathways
Bacteria have evolved regulatory systems that 1. couple expression of gene prod. to sensor systems 2. detect the amt of a particular comp. in the environment
Glucose levels control the lac operon Cyclic AMP= CAMP in bact +eukaryotes Camp= directly regulated by amt of glucose in environment. What = the role of camp in regulating the lac Operon?
CAP= Catabolite Activator Protein for Lac Operon Enzyme Adenelate Cyclase makes cAMP-it is directly regulated by glucose Cap alone will not activate anything-will not be activated to bind promoter on DNA Want lac operon to be off to use glucose Inverse reationship ↑ glucose- ↓ cAMP ↓ glucose- ↑ cAMP cAMP= needed to activate lac Operon will bind to Activator Protein Once cAMP binds CAP it binds promoter on DNA
Many DNA binding sites are symmetrical
DNA binding sites have rotational twofold symmetry
Allosteric effectors bind to regulatory proteins Allosteric effectors control the ability of activator or repressor proteins to bind to their DNA target sites. For many regulatory proteins, DNA binding is effected through the interaction of 2 different sites in the 3-dim structure of the protein. One site = is the DNA-binding domain. The other site, the allosteric site, acts as a sensor that sets the DNA-binding domain in one of two modes: functional or nonfunctional.
Effector Present Effector = Lactose that binds to protein (Activator) and changes it's shape bottom Effector= called Inducer when it binds repressor and pulls off repressor from DNA
Gene order in the trp operon corresponds to reaction order in the biosynthetic pathway The level of trp operon gene expression is governed by the level of tryptophan No tryptophan = trp gene expression is ↑ ↑ levels of tryptophan = trp operon is repressed
If too much Trp- can shut it down Tightly regulated essential AA-> turn on TRP Operator when TRP = not present b/c need > Ex. Arginine in Neurospora- 1 gene - 1 protein hypothesis 1st gene bottom precursor modified at each stage to become final AA-Tryptophan
Bact Attenuation: - Only in bact The trp mRNA leader sequence contains an - attenuator region- reg mech = built in -+2 tryptophan codons= TrpE gene
In the absence of the TrpR repressor protein, the presence of tryptophan halts transcription after the first 140 bases. In the absence of tryptophan, transcription of the operon continues.
Regulatory proteins control transcription
Repressor binds to operator-roadblock-obstructs promoter RNA binds to promoter What controls repressor from being bound? Lactose binds to repressor-changes it's shape-it comes off DNA-no more transcription
The lac operon is transcribed only in the presence of lactose
Repressor= constitutive protein-always made NO LACTOSE PRESENT don't need to make B-galactosidase RNA polymerase cannot get through LACTOSE PRESENT-acts as inducer takes repressor of DNA RNA polymerase makes repressor protein Lactose binds repressor-changes it's shape mRNA will have ribosomes transcribe DNA The relief of repression = called "induction" Lactose =called the "inducer"
Control of gene expression =goverened by DNA-binding proteins that recognize specific control sequences of genes
The Lac repressor protein binding to the lac operator
Diauxic growth of Escherichia coli on a mixture of glucose and lactose
Top: Lactose= present but B-galactosidase is not made. b/c glucose regulates the system called Catabolite Repression. pg 7 Catabolite Repression of the lac operon: Positive Control:2 Conditions needed in the environment: -Lactose must be present -Glucose must not be present
The 1st mechanism of regulation is just like The lac operon.... TrpR = the trp repressor TrpR binds tryptophan when tryptophan levels are ↑. In this situation, TrpR will bind to the operator + stop transcription.
When tryptophan was removed from the medium, trp mRNA was ↑ several-fold >
Negative and positive control of the lac operon HW: DRAW EACH SITUATION WITH EACH OF THE PROTEINS
b= in b/w position Lactose= Inducer when present- Repressor is off Operator No repressor + no activation there = very little lac mRNA c= No glucose present ↑ cAMP, lactose =present CAP-cAMP bound to promoter activator binding site Lactose Inducer binds Repressor + pulls it off
How do activators + repressors recognize when environmental conditions are appropriate for their actions and act accordingly?
for activator or repressor proteins to do their job, each must be able to exist in 2 seperate states: - 1 can bind to its DNA targets=DNA binding -1 cannot bind to its DNA targets The binding state must be appropriate to the set of physiological conditions present in the cell and its environment.