Flashcards in Lecture 8 Deck (18):
The lambda regulatory region:
- Contains PR (transcribes cro) and PL (transcribes N), which are transcribed by E.coli RNAP. They terminate at tR and tL
- cI, the repressor gene has it's own promoter called PRM (for repressor maintenance), expressed in the lysogenic pathway.
The right and left operators consist of...
and ____ and ____ bind proteins
- Three operator sites each with dyad symmetry, because each binds a dimer
- OR3, OR2 and OR1, which sit across PRM and PR.
- cI repressor protein and cro protein bind these operator sites.
Lambda repressor monomer:
- Has two structural domains that form an interacting dimer.
- Activating region
- Dimerisation region (monomers bind forming a dimer)
- Tetramerization region (dimer binding dimer)
Lambda cI dimers bind to an operator site cooperatively. This mean..
- cI first binds to OR1, then this binding enhances binding of cI to OR2.
- The repressor binds DNA via two alpha-helices in a dimer.
What does repressor do when it binds to OR1 and OR2?
- cI acts negatively to represses PR
- cI acts positively to activates PRM, by enhancing RNAP binding and creating cI gene
- Some cI mutations prevent positive regulation (the activating region) without affecting negative regulation (DNA binding)
cI acts to..
- Activate its own expression, while repressing cro expression, leading to repression of PR and PC and lysogeny.
- This is a positive feedback loop
cI also binds at OL1 and OL2 which can complex with
- cI at OR1 and OR2 (and OR3) in the lysogen.
- it only binds when the affinity is high
cro is a negative regulator that binds preferentially at OR3. It acts..
- negatively at PRM (overlaps OR3) repressing cI expression.
- as it increases cro binds at OR2 and OR1 (not cooperatively) repressing PR and it's own expression
During lysis cI and cro are..
- cI at OR2 and OR1
- PR off - no cro
- PRM on - more cI
- cI at OR3
- PRM off - regulates cl
During lysogeny cI and cro are..
- cro at OR3
- PRM off - no cI
- PR on - more cro
- cro at OR2 and/orOR1
- PR off, but PR is on, thanks to Q, so all the late cascade genes will be expressed
A plaque of WT lambda phage:
- means lysis
- it is turbid, so there are things (lysogens) growing in there.
A clear plaque is generated by a..
- cI mutant
What is involved in induction?
- Cleavage of cI by an E.coli protease (RecA).
- A lysogen going into the lytic cycle
- Acts as an anti-terminator of transcription to allow delayed early gene expression
- As the concentration of N protein builds up, more RNAPs will proceed to produced delayed early transcripts
How does anti-termination work?
- RNAP binds and transcribes N the anti-termination protein
- Anti-termination protein binds RNAP and prevents it from recognising RNAP, allowing it to pass the terminator
- Anti-termination proteins act on specific terminators.
- Termination is prevented by factors that act at nut, so rho cannot terminate.
- Are between promoters and terminators
Anti-terminators can act at different locations in the transcription unit eg):
- pN acts at nutL to enable RNAP to pass tL
- pN acts the nutR to enable RNAP to pass tR1
- pQ acts at put to enable rNAP to pass tR