10.4 - Lac Operon and PTS

Question 1

When glucose is present how is it important in LacY regulation?

Answer 1

Via membrane transporters IIC. Glucose will then move from IIC to IIB.

Question 2

What does IIB do when glucose is present in regulation of LacY?

Answer 2

IIB will transfer a phosphate from IIA to glucose, making G6P. The phosphate of IIA comes from HPr whose phosphate comes from PEP.

Question 3

What does the unphosphorylated IIA do when glucose is present in regulation of LacY?

Answer 3

The unphosphorylated IIA will bind and inhibit LacY (lactose permease).

Question 4

What happens when glucose is not present in regulation of LacY?

Answer 4

(1) When glucose is not present, the phosphate on IIA is not transported, therefore IIA is phosphorylated.
(2) The phosphorylated IIA cannot bind LacY.
(3) LacY is active and can import lactose.

Question 5

How is glucose imported when is is present in regulation of adenylate cyclase?

Answer 5

Glucose is imported via membrane transporters IIC. Glucose will then move from IIC to IIB.

Question 6

What does IIB do in regulation of adenylate cyclase?

Answer 6

IIB will transfer a phosphate from IIA to glucose. The phosphate of IIA comes from HPr whose phosphate comes from PEP.

Question 7

What does unphosphorylated IIA do in regulation of adenylate cyclase?

Answer 7

Unphosphorylated IIA binds adenylate cyclase and inhibits its activity. Adenylate cyclase is an enzyme required for cAMP formation. cAMP is not synthesized, it doesn’t bind CRP, CRP doesn’t bind DNA and transcription initiation of the lac operon cannot occur.

Question 8

What happens when glucose is not present in regulation of adenylate cyclase?

Answer 8

(1) The phosphate on IIA is not transferred, therefore IIA is phosphorylated.
(2) Phosphorylated IIA can no longer bind to adenylate cyclase, releases it and therefore activates it.
(3) Adenylate cyclase converts ATP to cAMP, which will bind CRP
(4) Conformational changes in CRP will allow it to bind operator sequence on DNA and initiate Lac operon transcription (and transcription of other operons).

Question 9

What is the sequence between the operator region and the actual genes?

Answer 9

The leader region

Question 10

What is the leader region transcribed into?

Answer 10

mRNA called leader mRNA

Question 11

How can leader mRNA fold?

Answer 11

(1) Pairing of regions 1 and 2.
(2) Pairing of regions 2 and 3 results in an anti-termination signal that allows transcription to continue.
(3) Pairing of regions 3 and 4 results in a termination signal which stops transcription further towards trp operon (Rho independent terminator).

Question 12

Why is transcription and translation coupled in the trp operon?

Answer 12

Because there is no nuclear membrane in bacteria

Question 13

What important sequence does leader mRNA contain?

Answer 13

Leader peptide coding region which contains sequence 1. Sequence 1 has two trp codons (very rare)

Question 14

What happens when there is not enough tryptophan in cells?

Answer 14

There are not enough tRNAs loaded with tryptophan so when the ribosomes get to these two trp codons on sequence 1, they will stall.

Question 15

What happens as a result of ribosomes stalling on sequence 1?

Answer 15

Sequences 2 and 3 will base pair. Sequences 1 and 2 cannot base pair because ribosomes are occupying sequence 1.

Question 16

What is the pairing of sequence 2 and 3 called?

Answer 16

This pairing is an antiterminator signal that allows RNA pol to proceed transcription. Transcription and then translation of the trp operon will occur and genes required for tryptophan synthesis are made.

Question 17

What happens when there is enough tryptophan in cells?

Answer 17

There would be enough tRNAs loaded with Trp. The ribosomes will read the 2 Trp codons on sequence 1 at a normal speed. Sequences 1-2 will form a loop and sequences 3-4 form another stem loop (Rho independent terminator).This Rho independent terminator will stop transcription by RNA pol and the trp operon doesn’t get transcribed.

Question 18

What is the stringent response?

Answer 18

A stress response of bacteria in reaction to amino acid starvation

Question 19

What happens when amino acids are scarce in regards to the stringent response?

Answer 19

Ribosomes will start stalling during translation because there are not a lot of loaded tRNAs.

Question 20

What does stallling in the stringent response lead to?

Answer 20

This stalling will activate RelA protein (pppGpp synthetase), which synthesizes ppGpp secondary messenger.

Question 21

What does ppGpp interact with in the stringent response?

Answer 21

ppGpp will interact with the \beta subunit of RNA polymerase leading to a decreased affinity to certain promoters (including rRNA gene promoters)

Question 22

What does ppGpp increase in the stringent response?

Answer 22

The affinity of RNA pol to other promoters of genes involved in responding to the amino acid station.