Theme 3- Module 2 (Prokaryotic transcriptional regulation) Flashcards
What two proteins increase in amount when E.coli cells transition from glucose to lactose metabolism?
Beta-galactosidase
Lactose permease
True or false: beta-galactosidase and lactose permease are detectable in small amounts when glucose is the primary energy source
False
They are not expressed until glucose is fully depleted from the growth medium
What is lactose permease?
Transmembrane protein that allows for the transport of lactose into the bacterial cells
What is beta-galactosidase?
Cytoplasmically situated bacterial enzyme that cleaves the imported lactose into glucose and galactose
What is a key advantage to the organization of the prokaryotic genome?
Groups of related genes with similar functions can often be found clustered together into operons. Leads to the ability to control the transcription of the whole gene cluster as one unit.
What do operons consist of?
A promoter
An operator (or on-off switch)
The coordinated gene cluster whose products will function in a common pathway or cellular response
What is an operator?
A sequence of nucleotides near the start of the operon that can allow or inhibit transcription
How do operators regulate transcription?
When the operator is not bound to any transcriptional inhibitor, then the RNA polymerase can attach to the promoter and transcribe the genes in the operon
What does the lac operon control in E.Coli?
The regulation of beta- galactosidase and lactose permease expression
What are the regulatory sequences of transcription in the lac operon?
- Promoter that binds the RNA polymerase complex
- Operator (lacO) which is the binding site for a repressor protein
What is the name of the sequence that the operator is expressed by?
Lac I coding sequence
What are the two main structural genes that code for the primary proteins needed to facilitate lactose metabolism?
lacY
lacZ
What does the lacY gene code for?
A lactose permease transport protein
What does the lacZ gene code for?
B-galactosidase
Which gene controls the expression of the lacZ and Y genes?
lac l
What does lac I code for?
A repressor protein which can bind to the operator and inhibit transcription from occurring
The ability of a repressor protein to halt transcription in this manner (the way that lac I halts transcription) is often identified as ________
Negative transcriptional regulation
How does negatively regulated transcription work?
Repressor protein will bind to the operator region of the operon
RNA polymerase is no longer able to bind to the promoter region of the operon
Results in turning off transcription
Is the lactose operon negatively or positively regulated?
Negatively
How does negatively regulated transcription work in the lac operon specifically?
Repressor protein, which is encoded by the lac l gene is constitutively expressed at low levels
(Repressor protein binds with the lac O operator region and the RNA polymerase complex is not able to bind to the promoter)
This is the state of the lac operon activity in most cases when E. coli cells are exposed to glucose
Describe the structure of the repressor in the lac operon
Tetrameric protein
- four identical protein subunits that binds tightly to operator regions on the lac operon DNA
How is the repressor able to prevent RNA Poly from binding?
Once all four subunits bind the lac operon DNA, that the DNA is twisted into a loop and RNA Poly is not able to bind
How does the presence of glucose in the growth medium affect the repressor?
Facilitates the constitutive expression of the repressor protein (and thus inhibits expression of lactase proteins)
How are repressor proteins allosterically inhibited by lactose?
Lactose = inducer molecule
- binds to the repressor proteins
- causes a change in the conformation of the repressor so that it can no longer bind to the DNA
What should you look out for when you’re looking at the general nutritional state of E. coli cells? (i.e. what’s a good indicator)
The conc of cAMP
When glucose levels are high in the environment surrounding bacterial cells, there is an inhibition of the enzyme __________
Adenylyl cyclase
What does adenylyl cyclase do?
Catalyzes the production of cAMP from ATP
When glucose levels are low, what will the conc of cAMP be like? Why?
High bc increased activity of the adenylyl cyclase enzyme
Describe the basic steps of positive regulation of the lac operon
1) Low levels of glucose –> high cAMP levels
2) cAMP binds to CRP (causes conformation change0
3) CRP-cAMP complex binds to CRP-cAMP binding site on DNA
3) activate transcription
If there was both glucose and lactose in the environment, would the CRP-cAMP complex bind to the lac operon?
No
Can transcription occur if the activator is not present or is not able to bind to the activator binding site?
No
What do inducer molecs do?
Allosterically inhibit repressor proteins so it can no longer bind to DNA
(bind and cause a conformation change)
What is the inducer molec involved in the lac operon?
Lactose (binds to the repressor)
