Chapter 20 - The Regulation of Gene Expression in Bacteria
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How does gene expression vary in bacteria?
varies with cell need.
Why do bacteria need to regulate gene expression?
to adapt to environmental changes quickly.
What has much of our knowledge about gene regulation come from?
studying gene regulation in bacteria.
Do bacteria use the same processes for gene regulation as eukaryotes?
No, bacteria do not use all the same processes as eukaryotes.
What type of gene regulation has most research focused on in bacteria?
Most of the research has been on transcriptional regulation.
What type of regulation has more information become available on recently?
translational regulation
What is the lac operon?
a set of genes involved in the metabolism of lactose.
What regulates the lac operon positively?
lactose.
What regulates the lac operon negatively?
glucose.
What are the genes included in the lac operon?
lacZ, lacY, and lacA.
What does the lacZ gene code for?
β-galactosidase.
What does the lac Y gene code for?
galactoside permease
What does the lac A gene code for?
Thiogalactoside transacetylase
What is the regulatory gene of the lac operon?
lacI, which encodes the lac repressor.
Is the lac I gene part of the lac operon?
No, the lacI gene is not part of the lac operon.
What type of enzymes does the lac operon code for?
catabolic enzymes.
When is the lac operon turned on?
presence of lactose and absence of glucose
What is allolactose?
Allolactose is the true inducer of the lactose operon.
How does allolactose interact with the lac repressor?
Allolactose binds to the lac repressor.
What is the relationship between lactose and allolactose?
Lactose is converted to allolactose by β-galactosidase through an alternative reaction.
What is the structure of lactose?
Lactose is galactose-(β1-4)-glucose.
What is the structure of allolactose?
Allolactose is galactose-(β1-6)-glucose.
What does the lac repressor bind to?
The operators
What type of protein is the lac repressor?
A homotetrameric protein with a helix-turn-helix domain
What does the lac repressor physically inhibit?
The binding of RNA polymerase
What is the length of the operator sequence in the lac operon?
21 base pairs
What is the major operator of the lac operon?
O1, which overlaps with the promoter
What are the auxiliary operators of the lac operon?
O2 (400 bp downstream) and O3 (90 bp upstream)
What happens to cAMP levels when glucose is low?
cAMP levels increase
What is cAMP derived from?
ATP
What does CAP bind to?
cAMP
What is the role of CAP:cAMP in gene regulation?
It is a global activator that can bind to more than 100 promoters
What is another name for the catabolite activator protein (CAP)?
cAMP receptor protein (CRP)
Where does CAP:cAMP bind in relation to the promoter?
Upstream at the CAP-binding site
What effect does CAP have on RNA polymerase and DNA interaction?
It helps stabilize the interaction
What happens to RNA polymerase without CAP?
It does not interact well with the DNA
What is the issue with the -35 sequence without CAP?
It is sub-par and lacks an UP-element
What is an inducer of the lac operon often used experimentally?
Isopropyl β-D-1-thiogalactopyranoside (IPTG)
Why is IPTG used in studies of the lac operon?
It cannot be metabolized, allowing researchers to study regulation without concern.
How does IPTG affect the lac operon?
It diffuses into the cell and binds to the lac repressor, turning the lac operon on.
What is a noninducer substrate of the lac operon?
X-gal
What does β-galactosidase do to X-gal?
It cleaves X-gal to produce galactose and 5-bromo-4-chloro-3-hydroxyindole.
What is produced when 5-bromo-4-chloro-3-hydroxyindole is oxidized?
5,5′-dibromo-4,4′-dichloro-indigo
What activates the promoter of the ara operon?
The presence of arabinose and absence of glucose.
What are the two activators required for the ara operon?
CAP and AraC
What happens to AraC in the absence of arabinose?
AraC binds as a homodimer to araI1 and araO2.
What occurs when arabinose is present?
Arabinose binds to AraC, forming a dimer that binds to araI1 and araI2.
What is required for the activation of the ara operon in the absence of glucose?
CAP:cAMP binds upstream of the araI site.
What is the role of a repressor in gene regulation?
It can hold RNA polymerase at the promoter.
Where does the Gal repressor (GalR) bind in E. coli?
To a site near the promoter, not overlapping.
What happens in the absence of galactose regarding GalR?
Gal repressor + RNA polymerase does not allow the closed to open complex.
What binds upstream of the gal site in the absence of glucose?
CAP:cAMP
What is the function of the merT operon?
It controls gene expression due to a physical change to the actual DNA.
How is MerR activated?
MerR is activated by mercury.
Does MerR interact with RNA polymerase?
No, MerR does not interact with RNA polymerase.
Where does MerR bind on the DNA?
MerR binds to a sequence between -10 and -35.
What is the distance between the -10 and -35 regions in the merT operon?
The distance is 19 bp, which is non-optimal for σ70.
What happens when MerR binds with Hg+2?
It causes a conformational change of the DNA, allowing RNA polymerase to bind.
What does the trp operon code for?
It codes for the five enzymes needed to make tryptophan.
What is the precursor that the trp operon converts to tryptophan?
The precursor is chorismate.
How does the trp operon respond to the presence of tryptophan?
It is turned off in the presence of tryptophan.
What type of enzymes does the trp operon code for?
It codes for anabolic enzymes.
What is a unique feature of the trp operon compared to the lac operon?
The trp operon lacks a CAP binding site.
What are the key components of the trp operon?
It contains an operator, promoter, leader region, and attenuator.
What is attenuation in the context of the trp operon?
It is an extra level of control that decreases transcription significantly.
How much does attenuation decrease transcription in the trp operon?
Attenuation decreases transcription by 10x.
What is the total decrease in transcription for the trp operon?
The total decrease is 700x.
Is attenuation present in the lac operon?
No, attenuation is not present in the lac operon.
In what type of cells does the trp operon function?
It works in cells without a nucleus, allowing coupled transcription/translation.
What is the leader sequence in the trp operon?
The leader sequence is trpL.
What is the promoter in the trp operon?
The promoter is trpP.
What is the operator in the trp operon?
The operator is trpO.
What is the effect of the absence of tryptophan (Trp) on the trp operon?
No trp repressor is present; inactive aporepressor is present.
What happens at low levels of tryptophan (Trp) in the trp operon?
No binding of tryptophan to aporepressor; no trp repressor is present.
What occurs in the presence of tryptophan (Trp) in the trp operon?
Tryptophan binds to aporepressor, leading to trp repressor binding to trp operator.
What is the role of tryptophan in the trp operon?
Tryptophan is the corepressor.
What is transcription attenuation in the context of the trp operon?
Regulation of bacterial operons by controlling termination of transcription at a site before the first structural gene.
Where is the attenuator located in the trp operon?
Within the leader sequence of the trp operon.
What forms the stem-and-loop structure in the trp operon?
Regions 3 and 4 of the leader sequence.
What is the probability of termination occurring in the trp operon?
There is a 90% chance of termination occurring.
What is the significance of the leader peptide in the trp operon?
It contains a pair of Trp codons that slow translation.
How many complementary sequences does the leader sequence have in the trp operon?
Four complementary sequences (1-4).
What are the two sets of mutually exclusive hairpins formed in the leader sequence?
Terminator (formed by 1•2 and 3•4) and anti-terminator (formed by 2•3).
What makes the terminator hairpin more stable than the anti-terminator hairpin?
The terminator has more base pairs.
What is the consequence of the ribosome stalling at the two Trp codons in the trp operon?
It prevents the formation of the 1•2 or 3•4 hairpins.
Why does the pairing choice in the leader sequence depend on translation?
Because transcription and translation occur simultaneously in prokaryotes.
What happens to the trp operon when tryptophan levels are high?
High [Trp] leads to ribosome translating the leader region until the stop codon between segments 1 and 2, causing segments 3 and 4 to interact and result in Rho-independent termination, preventing trp operon transcription.
What occurs to the trp operon when tryptophan levels are low?
Low [Trp] causes the ribosome to pause at the Trp codons in segment 1, allowing segment 2 to interact with segment 3 and form the hairpin (anti-terminator), leading to trp operon transcription.
What is the SOS response in cells?
The SOS response is the cell's reaction to DNA damage, which includes extensive breakage or mutation of the chromosome.
What proteins are involved in the SOS response?
The SOS response involves the LexA repressor protein and the RecA protein, as well as translesion synthesis (TLS) polymerases.
How does LexA affect the transcription of SOS genes?
LexA inhibits the transcription of SOS genes by binding to promoters.
What role does RecA play in the SOS response?
RecA helps with the autocleavage of LexA, leading to its dissociation and allowing the expression of SOS genes.
What is the role of ribosomal proteins in E. coli?
Ribosomal proteins regulate their own translation through feedback mechanisms.
How do encoded ribosomal proteins affect the ribosomal binding site (RBS)?
Encoded ribosomal proteins can bind to the RBS, inhibiting the small subunit from binding.
What is the relationship between free rRNA and ribosomal protein translation in E. coli?
The amount of free rRNA influences ribosomal protein translation; no free rRNA means no protein synthesis.
What happens when there is no free rRNA available for ribosomal proteins?
Ribosomal proteins cannot bind to the ribosomal binding site, leading to no translation.
What is the high-affinity binding behavior of ribosomal proteins in E. coli?
Ribosomal proteins bind to rRNA with high affinity when free rRNA is present.
What happens to ribosomal proteins when no rRNA is available?
Ribosomal proteins bind to a second highest affinity location on mRNA.
What type of bacteriophage is associated with E. coli?
Temperate λ phage.
What is a temperate bacteriophage?
A bacteriophage that can undergo both lytic and lysogenic cycles.