DD - Transcription II

Question 1

Constitutive genes vs regulated genes

Answer 1

Constitutive genes

• Housekeeping genes that are constantly expressed and needed for essential cellular functions

Regulated genes

• Genes whose expression is controlled and can be turned on or off in response to specific conditions or signals

Question 2

What are operons in bacteria?

Answer 2

Operons are:

• Units of gene expression, where genes encoding proteins in the same pathway are located together and transcribed into a polycistronic RNA.
• They lack introns.

Question 3

Where does control of transcription mainly occur?

Answer 3

At transcription initiation

Question 4

How are promoters recognised?

Answer 4

Promoters are recognised by RNA polymerase by having a consensus (common pattern of) DNA sequence

• hexamer (6bp) at -35 and a TATAAT sequence at -10
• asymmetric (only in 1 DNA strand), hence RNA polymerase knows which way to go

Question 5

What are down-mutations and up-mutations?

Answer 5

Down-mutations to decrease promoter efficiency usually decrease conformance to the consensus sequence

Up-mutations have the opposite effect

Question 6

What is Bacterial RNA polymerase made up of?

Answer 6

Bacterial RNA Polymerase

Holoenzyme (complete enzyme) consists of 5 types of subunit (6 units)

2 ɑ subunits (40 kD) – enzyme assembly

β, β’ (prime) = form catalytic centre

ω subunit – enzyme assembly and stability

σ sigma (70 kD) – binds promoter

Question 7

What is a regulon?

Answer 7

A regulon is a group of genes that are regulated as a unit

Question 8

What are the 2 types of transcriptional regulators?

Answer 8

Negative regulation - transcriptional repressors

• Repressor binds to sites known as Operator sites
• Stops RNA polymerase binding

Positive regulation - Transcriptional activators

• Activator binds to specific site
• Helps RNA polymerase bind

Question 9

What is the negative and positive regulator for the lac operon?

Answer 9

Negative regulator- Lac repressor (lacl)

Positive regulator- Catabolite activating protein (CAP)

Question 10

What are the 3 structural genes present in the lac operon and what do they do?

Answer 10

β Galactosidase cleaves lactose into its component sugars

Permease transports lactose into cells

Transacetylase covalently modifies lactose

Question 11

What occurs when lactose is absent and present?

Answer 11

Lactose absent

• Lac repressor is produced
• Lac repressor binds to the operator
• Lac operon transcription blocked

Lactose present

• Allolactose (derived from lactose metabolism) is produced
• Allolactose binds to the lac repressor
• This induces a conformational change in LacI
• Repressor cannot bind to operator

Question 12

What are features of the lac repressor? (3)

Answer 12

1) Is a tetramer

2) Contains 2 dimers

• Each dimer within the tetramer can bind to one operator site

3) Must bind to 2 out of 3 sites

• Repressor can bind to O1 and O2 or O1 and O3
• But not to O2 and O3

Question 13

What occurs when there is a low level of glucose?

Answer 13

Low Glucose → cAMP high → CAP active → Lac Operon active → Lactose breakdown

Question 14

What causes termination of transcription?

Answer 14

A run of A-Ts in the template strand

Question 15

What does the stem-loop. structure in the RNA cause?

Answer 15

Causes RNA polymerase to pause → DNA hybrid unravels from the weakly bonded A

Question 16

What are properties of Rho?

Answer 16

1) Prokaryotic transcription protein

2) ~275kD hexamer

3) Each subunit has a:

• RNA binding domain
• ATP hydrolysis domain: moves along the RNA

4) Stalls the RNA polymerase

5) Rho is a helicase

• Unwinds the DNA:RNA hybrid