DNA Replication #2

Question 1

Name four different methods of bacterial regulation fo initiation.

Answer 1

1. Regulatory inactivation of DnaA
2. Mehtylation
3. DnaA sequestration
4. Recycling of DnaA-ADP for DnaA-ATP

Question 2

Caf1

Answer 2

Chromatin assembly factor 1

~ mediates chromatin assembly in DNA replication and DNA repair

Question 3

SeqA

Answer 3

binds to hemi-methylated GATC sites and prevents re-methylation/binding of DnaA
~ only a temporary block - dissociates then process continues unless another SeqA binds

Question 4

RIDA

Answer 4

Regulatory inactivation of DnaA (RIDA)
~ blocks replication initiation in bacteria
~ accelerates active ATP-DnaA being converted to the inactive ADP-bound form

Question 5

Reverse transcriptase

Answer 5

~ DNA pols that copy RNA into DNA

~ ssRNA =template

Question 6

Telomerase

Answer 6

specialised reverse transcriptase

~ made up of protein (TERT) and RNA (template for repeats + role in enzyme activity)

Question 7

Telomeres

Answer 7

simple sequence repeats at the ends of chromosomes to prevent loss of information
~ TTAGGG in humans
~ G-rich

Question 8

The End-repliclation problem

Answer 8

lagging strand synthesis cannot replicate the very end of a linear chromosome
~ not a problem for circular chromosomes

Question 9

What promotes Okazaki fragment maturation in eukaryotes?

Answer 9

Flap endonuclease (Fen1)

Question 10

What promotes Okazaki fragment maturation in E.coli?

Answer 10

DNA pol 1

Question 11

Polymerase switching (eukaryotes)

Answer 11

handover from Pol alpha-primase to pol delta or epsilon

~ facilitated by clamp loading

Question 12

How does DNA ligase work?

Answer 12

~ nucleophilic attack
~ requires nucleotide cofactors:
BACTERIA = NAD+
EUKARYOTES = ATP

Question 13

What are the clamp loaders in bacteria/eukaryotes?

Answer 13

Bacteria = gamma complex
Eukaryotes = Replication factor C (RFC)

Question 14

What are the sliding clamps in bacteria/eukaryotes?

Answer 14

Bacteria = B-protein
Eukaryotes = PCNA

Question 15

Name the indicator proteins in bacteria/eukaryotes.

Answer 15

Bacteria = DnaA
Eukaryotes = ORC

Question 16

What are the bacteria/eukaryotes DNA primases called?

Answer 16

Bacteria = DnaG
Eukaryotes = polymerase-alpha-primase

Question 17

Termination in bacteria DNA replication

Answer 17

~ ter site is bound by Tus
~ replication stops
~ DNA pol I fills in un-replicated DNA

Question 18

Name 4 regulatory mechanisms for DNA rep. initiation used in bacteria.

Answer 18

~ regulatory inactivation of DnaA (RIDA)
~ methylation of GATC sites by DNA adenine methylase
~ DnaA sequestration (its binds to datA region next to orC)
~ Recycling of DnaA-ADP to ATP (binds to DARS region and acts as a cofactor for the exchange)

Question 19

Name … regulatory mechanisms for DNA rep. initiation used in bacteria.

Answer 19

~ formation of pre-replication complex (PreRC)
~ S-phase kinases phosphorylate MCM, Cdc6 + Cdt1 = released and degraded
~ Cdc45-MCM-GINS complex (CMG) unwinds helix at each fork
~ effect of Cdk levels

Question 20

Pre-replication complex (PreRC)

Answer 20

~ initiates DNA rep but then inactivates components

~ only forms in G1 as low Cdk levels

Question 21

Regulatory effects of Cdk levels

Answer 21

S1 = decrease cdk lvls = allows formation of preRC
G1 = increased cdk lvls = activates preRC