The Molecular Process of DNA Replication

Question 1

How accurate is DNA polymerase?

Answer 1

It only makes 1 error in every 10 million nucleotide pairs it copies

Question 2

How is DNA polymerase so accurate?

Answer 2

It has error correcting activity -> proofreading

Question 3

What does DNA pol do before adding a new nt?

Answer 3

It checks the previous nt to see if its correct

Question 4

What happens if DNA pol finds out the previous nt is incorrect?
(3)

Answer 4

DNA pol removes the incorrect nt by cutting the phosphodiester bond

This releases the nt

DNA pol tries again to add the correct nt

Question 5

What is DNA polymerase III?

Answer 5

The main replicating enzyme in bacteria

Question 6

What does DNA polymerase III do?

2

Answer 6

DNA polymerase activity in forward direction

Exonuclease activity in reverse - proofreading

Question 7

What does DNA polymerase I do?

Answer 7

Replaces RNA in primer with DNA

In the forward direction exonuclease activity mediating nick translation during DNA repair

Question 8

What does DNA polymerase I do in the forward direction?

2

Answer 8

DNA polymerase activity

Exonuclease activity - nick translation during DNA repair

Question 9

What does DNA polymerase III do in the reverse direction?

Answer 9

Exonuclease activity - proofreading

Question 10

What is an exonuclease?

Answer 10

An enzyme which removes successive nucleotides from the end of a polynucleotide molecule

Question 11

What are the four key players in DNA replication?

Answer 11

Ligase

Helicase

Primase

DNA polymerase

Question 12

What does Primase do?

Answer 12

Adds the RNA primers

Question 13

What does helicase do?

Answer 13

Unzips the helix

Question 14

What does ligase do?

Answer 14

Glues the Okazaki fragments together

Question 15

What needs to be done to start a new DNA strand?

Answer 15

Primase adds short segments (5-10 nts) of RNA bases first to provide a base-paired 3’ end as a starting point for DNA pol

Question 16

How many primers are needed on the leading strand?

Answer 16

Only one

Question 17

How many primers are needed on the lagging strand?

Answer 17

New primers are needed continuously

Question 18

What three additional enzymes are needed on the lagging strand?

Answer 18

Nuclease

DNA polymerase I (bacteria only)

DNA ligase

Question 19

What does nuclease do?

Answer 19

Breaks apart the RNA primer

Question 20

What does DNA polymerase I do in bacteria?

Answer 20

Replaces RNA with DNA

Question 21

What does DNA ligase do?

Answer 21

Joins 5’ phosphate end of one DNA fragment to 3’ OH end of the next (ATP required for ligase activity)

Question 22

What four proteins cooperate to form the replication machine?

Answer 22

Helicase

Topoisomerase

Sliding clamp

Single strand binding proteins

Question 23

Write a note on helicase.

3

Answer 23

An enzyme

Uses energy of ATP hydrolysis to speed along DNA and separate the strands of parental DNA ahead of polymerase

It breaks the hydrogen bonds between base pairs

Question 24

Write a note on topoisomerase.

2

Answer 24

An enzyme

Prevents DNA helix upstream from becoming supercoiled by relaxing it

Question 25

Write a note on a sliding clamp.

2

Answer 25

Keeps DNA polymerase firmly attached to DNA template

On the lagging strand it releases polymerase from the DNA each time an Okazaki fragment is completed