Wright 8 - Catabolite Operons and Regulatory Proteins

Question 1

What happens to lactose metabolism enzyme activity when lactose is added? When glucose is added? What is this due to?

Answer 1

Lactose: Increases
Glucose: Decreases

Due to catabolite repression (the glucose effect)
- This can be observed with any sugar (not just lactose)

Question 2

For these four carbon sources, when is growth rate of E. coli. fastest?

• Glucose
• Lactose
• Arabinose
• Glycerol

Answer 2

Glucose, 30 minutes.

Question 3

For these four carbon sources, which ones have low/high cAMP levels? Why?

• Glucose
• Lactose
• Arabinose
• Glycerol

Answer 3

Glucose: low
Lactose: high
Arabinose: high
Glycerol: high

Glucose inhibits the activity of the enzyme adenylate cyclase, thereby, decreasing the intracellular concentration of cAMP in the bacterial cell when glucose is present.

Question 4

What is the catabolite activator protein (CAP) required for?

Answer 4

Maximal initiation of the lactose operon and other operons that code for enzyme required for sugar metabolism in E. coli. It binds to the CAP site in the promoter. This causes the entry site for RNA polymerase to become destabilized and allow RNA polymerase binding.

Question 5

How is transcription terminated rho-independently?

Answer 5

1. A rho-independent terminator contains an inverted repeat followed by string of approximately six adenine nucleotides
2. The inverted repeats are transcribed into RNA
3. The inverted repeats in RNA fold into a hairpin loop
4. The hairpin loop causes the RNA polymerase to stop
5. There is a string of uracils after the hairpin, these allow DNA-RNA pairing to become destabilized and cleaved

Question 6

How is transcription terminated by the rho factor in a cell-free in vitro system?

Answer 6

There are rho-dependent terminators along a DNA template, to transcription is terminated at different sites depending on when rho factor is added

RNA polymerase pauses when it encounters a terminator sequence, rho catches up and unwinds the RNA-DNA hybrid and brings transcription to an end

Question 7

What are the differences between rho-independent and rho-dependent termination in terms of the stem structure?

Answer 7

Rho-independent

• GC-rich stem
• Run of uracils after

Rho-dependent

• No GC rich stem
• No run of uracils

Question 8

What does concentration of rho factor cause?

Answer 8

Depending on the abundance of rho, different mRNAs are produced, generating different polypeptides encoded by a single operon.