phage development

Question 1

what is bacteriophage lambda

Answer 1

it is a phage which predates E. coli. it has 20 faces and encodes around 50 genes on 7 operons

Question 2

what is the lytic phase

Answer 2

active replication where around 100 new phage are produced and the E.coli host is lysed

Question 3

what is the lysogenic phase

Answer 3

integration of the phage genome into the host. it can re-enter the lytic cycle under the influence of UV

Question 4

which phase is favoured by which environment

Answer 4

is rapidly growing E.coli the lytic phase is favoured, in weak E.coli populations the lysogenic is favoured as the bacteria can’t support a growing phage population

Question 5

why does DNA circularise upon injection in the lytic cycle

Answer 5

to protect it from host exonucleases. it also brings together certain genes for coordinated expression

Question 6

what part of the phage genome is transcribed first, in the very early stage, and by what

Answer 6

the N and Cro regions are transcribed first by PL and PR promoters, and are terminated by tL1 and tP1 respectively

Question 7

how does transcription take place beyone the tL1 and tP1 terminators

Answer 7

the N protein is an anti-terminator and alters the RNAP in Nut sites

Question 8

what is transcribed by PL and PR in the early stage

Answer 8

PL transcribes CIII, xis and int and PR transcribes CII, O, P and Q

Question 9

what is the role of the Q protein

Answer 9

it modifies RNAP at the Qut sites to allow transcription of the head and tail regions of the genome in the late stage

Question 10

what is the role of Cro in the late stage

Answer 10

it prevents the synthesis of CI (a repressor) and further transcription of early stage genes

Question 11

which new promoters are available to bind RNAP in the lysogenic pathway

Answer 11

Pint for the transcription of the int gene and PRE for the transcription of CI

Question 12

which genes are required for integration and excision

Answer 12

int is needed for integration and both int and xis are needed for excision

Question 13

what is the role of CII and when is it most abundant

Answer 13

it stimulates the production of int. it is protected from degradation by CIII and is most abundant when the host proteases are low which is during periods of starvation

Question 14

how is int regulated in the lytic phase

Answer 14

both int and xis are transcribed, but so is sib which degrades int mRNA so levels of int are lower relative to xis

Question 15

what promotes the CI gene for the lambda repressor

Answer 15

it is initially transcribed by the PRE promoter, then later by the PRM promoter

Question 16

what differences are there in the affinity of the lambda repressor and Cro proteins

Answer 16

lambda repressor has high affinity for OR1, and as a result of being a dimer OR2, whereas Cro proteins have a higher affinity for OR3

Question 17

how does the lambda repressor positively and negatively regulate simultaniously

Answer 17

by binding to OR1/2 it stimulates PRM and therefore increased the amount of transcription of CI to produce more of the lambda repressor (+). this simultaneously blocks the PR promoter which is needed for the transcription of the Cro and other lytic genes (-)

Question 18

what is the result of lambda repressor self-regulation

Answer 18

it allows the repressor to be made in excess which is used to prevent super infection

Question 19

describe the structure of the lambda repressor

Answer 19

each monomer has two domains, an amino and a carboxyl, but exists as a dimer

Question 20

what are the roles of the two domains of the lambda repressor

Answer 20

the C domain is used to bind to another monomer to form a dimer. the N domain is what binds to the DNA in the OR operon

Question 21

when will Cro proteins bind each of the OR operon domains

Answer 21

it will initially bind to OR3 as it has highest affinity for this. it will only bind OR1/2 when it is at a very high concentration as this blocks the transcription of more Cro genes

Question 22

how do the lambda repressor and Cro proteins bind to the DNA of the OR operon

Answer 22

the DNA domains have two- fold mirror symmetry and two major grooves- one for each monomer of the dimer to bind

Question 23

what do both the lambda repressor and Cro proteins use to bind to the OR DNA

Answer 23

they both use a helix- turn- helix motif

Question 24

what determines the affinity of Cro and lambda repressor proteins for the OR DNA

Answer 24

differences in the amino acid- base pain interactions across the three OR domains

Question 25

what is a lysogen

Answer 25

when the lambda repressor genome has been integrated into the bacterial chromosome

Question 26

what is the significance of the CI gene in regards to inducing the lysogen

Answer 26

the CI gene is the only one transcribed in the late stages of the lysogenic pathway. it is the deactivation of CI that induces the lysogen to enter the lytic pathway

Question 27

what is RecA

Answer 27

a host DNA repair protein which is activated by UV light

Question 28

how does RecA deactivate CI

Answer 28

it cleaves the bond between the C and N domains of the lambda repressor so they can’t dimerise and lose affinity for the OR operon domains. PRM isn’t stimulated and PR isn’t blocked

Question 29

what is the normal function of RecA and how is it exploited by the lambda phage

Answer 29

it is normally part of the bacteria’s SOS response to cleave bacterial protein LexA. the phage exploits it due to the structural similarity of LexA and the lambda repressor

Question 30

how does excision happen in the induction of the lysogen

Answer 30

due to the transcription of equal amounts of the xis and int genes

Question 31

why is sib not transcribed in the induction of the lysogen

Answer 31

it is only brought close to the int/xis genes when the genome circularises upon entering the lytic cycle