Lecture 6: Bacterial Genome Replication and Regulation 2 Flashcards
What is a constitutive gene?
-gene that is always expressed
What is an inducible gene?
- gene that requires activation. is normally in the off state
What is a repressible gene?
-gene that is normally on, and is able to be turned off.
The beta-galactosidase enzyme is what type of regulation?
inducible
The Trp operon utilizes what type of regulation?
- repressible
How do regulatory proteins control transcription initiation?
- contain DNA binding domains in order to interact with DNA
- inhibit or promote transcription
- utilize allosteric regulatory sites (coreceptors)
What happens to induce negative control of an inducible gene?
-a repressor protein binds at the operator, and prevents binding of RNA polymerase and therefore prevents transcription
What control is being applied if a repressor protein is bound at a promoter but no transcription is taking place?
negative control of inducible gene
How can an inducible gene under negative control start transcription?
the inducer binds with the repressor protein, removing it from the promoter and allowing transcription to occur
Negative control of a repressible gene with no presence of a co-repressor produces what effect?
without the co-repressor the repressor protein will not bind to the promoter and transcription will occur
Negative control of a repressible gene with the presence of a co-repressor produces what effect?
the co-repressor will bind with the repressor protein, and then bind the promoter and inhibit transcription
Positive control of an inducible gene with no inducer has what effect?
no inducer is able to activate the activator protein, which is required to start transcription.
-transcription will not take place
Positive control of an inducible gene when the inducer is present has what effect?
the inducer binds to the activator protein and binds to the promoter region, which will activate transcription
Positive control of a repressible gene when there is no inhibitor present has what effect?
no inhibitor, means the activator protein is bound and allows transcription to take place
Positive control of a repressible gene when there is an inhibitor present will have what effects?
the inhibitor will bind with the activator gene and prevent binding at the promoter region. Thus preventing transcription
What happens when the lac repressor binds to the operator?
The repressor causes a conformational change of the lac operator, by looping it, to conceal/hide the coding sequence
When does the lac repressor become active? Or when does it bind to the lac operon?
lac repressor binds to the lac operon and prevents transcription when lactose is not present
When lactose is present in an environment what happens to the lac operon?
- lacl (regulatory region) transcribes mRNA to form allolactose which binds the lac repressor
- inhibited lac repressor canât bind to lac operator.
- RNA polymerase transcribes the lac operon
The use of the lac repressor gene is characteristic of what type of gene control?
negative
Positive control of the lac operon utilizes what proteins?
CAP; catabolite activator protein, which requires cAMP as a co-activator
What does the presence of glucose do to the CAP?
-glucose presence prevents binding of CAP, and therefore prevents transcription
What happens to CAP in the absence of glucose?
with no glucose present the CAP is activated and transcription can occur depending on the repressor status
What happens to the lac operon when there is lactose but no glucose present?
- no glucose activates cAMP to bind with CAP, causing it to bind to the CAP site. (activate transcription)
- presence of lactose will produce allolactose, which will prevent the repressor from binding
- activator is active, repressor is not bound to operatorâ> transcription occurs
What happens to lac operon when neither lactose of glucose are present?
- no glucose activates cAMP to activate CAP. (activates transcription)
- no lactose, causes lac repressor to bind to lac promoter/operator (inhibit transcription)
- no transcription will occur
What happens when lactose and glucose are present?
- glucose presence prevents cAMP forming, prevents activation of CAP (no transcription)
- lactose presence binds the lac repressor with allolactose (promoting transcription)
- no activator proteinâ> no transcription
What happens when glucose is present but no lactose is present?
- glucose presence prevents cAMP forming, preventing activation of CAP (no transcription)
- no lactose allows repressor to bind lac operator and promoter (no transcription)
- no transcription occurs at all
Why are the levels of cAMP based on the level of glucose?
the catalysis of PEP produces phosphate that must go somewhere.
- glucose is present: phosphate is transferred to glucose;cAMP levels decline
- no glucose; phosphate transferred to adenyl cyclase, which activates cAMP
The Trp operon is mostly under what type of control?
- negative control via repressible genes
What happens with the Trp operon during low levels of tryptophan?
- The trpR codes for mRNA to produce an inactive trp repressor.
- Low levels of trp, will not have any trp available to bind to the repressor.
- RNA polym will bind to the promoter and transcription will begin
What happens to the trp operon when trp is readily available within the body?
- The trpR codes and forms mRNA to form the repressible protein.
- Excess trp is readily able to bind to the repressor protein
- The repressor then binds to the operator and prevents transcription of trp.
Why does it make sense for the trp operon to be under negative repressible control?
If the body already has readily available sources of trp, it does not need to waste energy to form more. Therefore the gene is âhiddenâ to conserve energy
What happens to the ara operon when arabinose is not present in the cell?
- Arabinose will not be available to bind to the AraC protein
- The AraC protein will form a dimer between the binding sites of the operator and the initiation site.
- This will prevent binding of RNA polymerase, and arabinose will not be produced.
What happens to the ara operon when arabinose is readily available to the cell?
- Arabinose binds with the AraC protein, breaking the dimer loop.
- The AraC proteins form dimers but on the same motif, therefore not causing a structural alteration to the DNA strand
- CAP+cAMP protein is able to bind to promote transcription
- RNA polymerase binds and begins transcription
What is attenuation?
- The controlled regulation of termination of transcription
Riboswitches play a large role in what function?
- contributing to attenuation regulation
Attenuation of the trp operon leads to the formation of what structures?
Hairpins form on the RNA strand that control the transcription.
-this is controlled by what regions of the RNA are pairing together to form hairpins
What occurs if hairpin is formed at 1:2 and 3:4 of the trp operon RNA?
No translation occurs. 3:4 pairing indicates high levels of trp in the cell
If regions 2:3 form a hairpin in trp operon RNA what is the resulting outcome?
1.regions 2:3 form a hairpin due to low levels of trp, therefore translation will occur to produce trp
If region 3:4 are able to form a hairpin what is the resutling outcome, in trp operon?
- Translation occurs
- At this point the 3:4 hairpin forms in response to high levels of Trp.
- High Trp levels stop the transcription of trp operon.
What is the riboswitch similar to ?
- the mRNA leader sequence
How do riboswitches control transcription?
- riboswitches alter the folding pattern, with stimulation from the proper effector, of the mRNA leader sequence
What bacteria have been found to have translation inhibited by riboswitches?
Gram-negative bacteria
A riboswitch is a ligand that binds to DNA or mRNA to inhibit transcription or translation. If a riboswitch wishes to inhibit translation of a gram-negative bacteria, what coding sequence must be âhiddenâ from the ribosome?
- shine dalgarno sequence
- - binding site for the ribosome, which is just upstream of the start codon.
What is the function of small RNA (sRNA) or noncoding RNA (ncRNA)?
- Some are complementary to mRNA and can base pair
2. They can inhibit or enhance translation
Why do bacteria use global regulatory systems such as regulons and modulons?
- Allows them to respond rapidly to a variety of changing conditions
What is a regulon?
- Gene or operon controlled by a common regulatory protein
What is a modulon?
- an operon network under control of a common global regulatory protein
What is an example of modulons?
- the lac operon and CAP proteins
What mechanisms does global regulation incorporate to provide regulation?
- Two component signal transduction system
- Phosphorelay system
- Regulatory proteins
- Alternative sigma factors
The 2 component regulatory system is found where, and generally uses what two proteins?
- Archaea, eukaryotes, prokaryotes
- Sensor kinase
- Response-regulator protein that is activated by the sensor kinase.
What is the function of the sensor kinase in the two component regulatory system?
- Act as extracellular receptor for metabolites
2. Relates to intracellular communication pathway
What is the function of the response-regulator protein in the 2-component regulatory system?
- Activated by the sensor kinase
2. Binds to DNA to activate or inhibit transcription
What is Env Z?
- sensor kinase that autophosphorylates in high osmolarity
What is OmpR?
- the promoter region that controls the formation of ompC or ompF
What regulates expression of porin proteins based on osmolarity?
OmpC and OmpF
What is the function of the porin protein OmpC?
- small porin protein
- expressed with high E.coli osmolarity
- will lower level of diffusion due to smaller size
What effect does the expression of OmpF, porin protein, on E.coli?
-it will be expressed when in a dilute environment and large amount of solutes will need to diffuse
What happens in E. coli to cause the pathogen to stop running and move towards a chemotactic material? Would occur with high levels of chemotactic in spots
- Methyl-accepting proteins are methylated
- CheA/W are phosphorylated
- CheY is active and causes basal body to produce clockwise rotation
What happens to E. Coli to produce the running effect from a chemotactic material? Would be in low levels of chemotactic.
- MCP (methyl-associated chemotactic protein) is not methylated
- CheA/W are not active
- CheY is not phosphorylated
- Basal body produces CCW rotation with use of proton gradient.
What is quorum sensing?
> cell-to-cell communication mediated by small signaling molecules such as N-acyl-homoserine lactone (AHL)
If envZ is stimulated to phosphorylate ompR what happens?
-ompR phosphorylated will inhibit formation of ompF and promote formation of ompC
If envZ is not stimulated, ompR will not be phosphorylated. What is the overall reaction?
-unphosphorylated ompR will lead to inhibition of the ompC, and formation of the ompR
What regulator is in charge of transcription of proteins for fluroescence in the V.fisheri?
- LuxR regulator that activates transcription
What process will produce light production in the V.fisheri?
- AHL must be produced in high level and concentration
- These diffuse into the cell, bind and activate LuxR.
- LuxR stimulates AHL synthase (luxl) and produces proteins
- products produce light
What happens in V.fisheri and other similar organisms if there is low cell density? (low AHL)
- low autoinducer present
2. LuxR is not made and no light produced
What happens in V.fisheri and other organisms if cell density of AHL is high?
- increases combination ability of autoinducers
2. LuxR is activated and produces luminescence
When will TTSS inhibit the process of bioluminescence and when will it promote bioluminescence?
- low cell densities; too few autoinducers to inhibit kinase activity and LuxR not formed, and TTSS not blocked
- high cell densities; kinase activity promoted an LuxR formed to block the TTSS
The stringent response in E.Coli occurs when, and what happens?
- occurs: with unequal amounts of tRNA and AA.
2. Complex RelA binds to the ribosome and catalyzes the reaction of GDP to guanine tetraphosphate
What effects will guanine tetraphosphate have when combined with DksA, on RNA polymerase?
- will downregulate gene expression if sigma is at GC rich region
- will upregulate gene expression if sigma is at TA rich region
During bacillus sporulation, what is the effect of having an active Kin A/B?
- this will phosphorylate and ACTIVATE the cascade sequence leading to early and late sporulation gene transcription
How are induce mutations caused, and what are common types?
- any agent that directly damages DNA
2. can form base analogues, intercalating agent, alkylating agents
What is a forward mutation?
the mutation of a wild type into a mutant form
What is reverse mutation?
a mutant phenotype converts back to a wild type phenotype
- is common with supressor genes
What is an auxotrophic mutant and where do they come from?
- mutant unable to make essentials for survival
2. from a wild-type prototroph
What is the role of RecA in the SOS Response?
- initiate recombination repair
2. protease, destroys LexA repressor, which increases excision repair enzyme
With respect to the SOS Response, what synthesize unrepaired DNA?
DNA pol. IV, V
How does horizontal gene transfer differ from vertical gene transfer?
there is no generation skipping, and genes are transferred from a mature organism to another.
Why is horizontal gene transfer useful?
- increases virulence
- gene transfer between same or different species
- transformation, conjugation, transduction
What processes of donor DNA and recipient chromosome combination will produce stable recombinants of the two DNA?
- integrate donor DNA into the recipient DNA, and undergo reproduction
- Donor DNA can self replicate, as a plasmid, and live together with the recipient DNA
What process of donor DNA and recipient DNA recombination occurs to produce non-stable recombinants?
- Donor DNA unable to replicate, and host DNA able to reproduce; eventually dies off
- Host DNA can attack the donor, forming peptide fragments; then dying off after reproduction.
What results in Fâ conjugation?
- the F factor incorrectly leaves the host, and leaves some behind
- causes some of host genes to be removed with some of F factor
Can genes be conjugated after Fâ conjugation?
yes
What size material is able to be taken up by bacteria via transformation?
- single strand of DNA
- which then aligns itself to homologous region
- homologous regions bind and form heteroduplex DNA
What is transduction?
- transfer of bacterial genes via a virus
What two ways can viral DNA invade a bacteria via transduction?
- lytic or lysogenic cycles
What is unique to generalized transduction?
- transfer of any part of genome
- occurs in virulent lytic cycle
- DNA host fragments are mistakenly packed into phage head
What is able to perform specialized transduction?
- temperate phages that establish lysogeny
What parts of the genome are transferred in specialized transduction?
- specific portions
- only occurs when prophage is incorrectly excised
When are donor genes and not F factor generally transferred to a recipient?
during HFr conjugation
How is the plasmid replicated in F+ x F- mating?
the plasmid replicates by the rolling circle method
