Enzymes in DNA replication

Question 1

In Ecoli- What does DNA polymerase 1 do? What 3 activities?

Answer 1

1) DNA synthesis through polymerisation of nucleotides to a free 3’OH
2) 3’ to 5’ exonuclease activity - important for proofreading newly synthesised DNA to ensure if enzyme inserts incorrect nucleotide, it can be removed and replaced w correct one.
3) 5’ to 3’ exonuclease to degrade a strand ahead of the advancing polymerase such as the laid down RNA primers and in DNA repair processes.

Question 2

In Ecoli - Where are these activities of polymerase 1 found on the enzyme

Answer 2

-The first 2 activitiues (polymerisation and 3to5 exonuclease activity) are found on a large domain of the enzyme called the Klenow fragment
-Other activity (5to3 exonuclease activity)
found on a smaller fragment

Question 3

In Ecoli - What encodes DNA polymerase 1?

Answer 3

polA gene

Question 4

In Ecoli - What are DNA polymerase 1 and 2 involved in and what is DNA polymerase 3 responsible for?

Answer 4

DNA polymerase 1 and 2 are involved in DNA repair
DNA polymerase 3 is the enzyme responsible for bacterial DNA replication

Question 5

DNA polymerase 3 of Ecoli - 3 subunits that comprie the core of enzyme functions?

Answer 5

-alpha - has the DNA polymerase activity
-e - has a 3’ to 5’ exonuclease that removes mismatched bases as soon as they are incorporated
-0 subunit stimulates the e subunits proofreading

Question 6

Eukaryotic DNA polymerases subunits

Answer 6

-Multiple DNA polymerases participate
Replication:
alpha- priming synthesis
delta- elongating lagging strands
e- leading strand replication

ß- repair
Y- may be involved in replicating mitochondrial DNA

Question 7

What does helicase do?

Answer 7

Uses energy from ATP to separate parental strands

Question 8

What does Primase do?

Answer 8

Binds to helicase to form a primosome

Question 9

What is Primase activated by?

Answer 9

DNA helicase, then synthesises a short RNA primer approx 11 nucleotides long to which new nucleotides can bve added by DNA polymerase

Question 10

What do topoisomerase 1 and DNA gyrase do?

Answer 10

Wind and unwind supercoiled DNA to facilitate DNA replication

Question 11

What does single strand DNA binding protein do?

Answer 11

Bind to separated DNA strands, coat them so they cannot re-anneal