DNA replication in prokaryotes

Question 1

What is the replication origin?

Answer 1

The point on a DNA sequence where replication begins

Question 2

What is the main DNA replicating enzyme in prokaryotes?

Answer 2

DNA polymerase III

Question 3

What is the function of DNA polymerase I in prokaryotes?

Answer 3

DNA replication and repair

Question 4

What is the function of DNA polymerase II in prokaryotes?

Answer 4

DNA repair

Question 5

What is the function of DNA polymerase VI and V in prokaryotes?

Answer 5

DNA repair

Question 6

What is the shape of DNA polymerase referred to as?

Answer 6

A hand, containing fingers, a thumb, and a palm

Question 7

Where on DNA polymerase does the catalytic activity occur?

Answer 7

The palm

Question 8

How many protein subunits is DNA polymerase III made up of in prokaryotes?

Answer 8

9

Question 9

Does DNA polymerase III in prokaryotes have 3’ to 5’ exonuclease activity?

Answer 9

Yes

Question 10

What does 3’ to 5’ exonuclease activity mean?

Answer 10

The enzyme can proofread the copy being made to minimise copying mistakes

Question 11

What cofactor is needed for DNA polymerase to work?

Answer 11

Mg²⁺

Question 12

What are the 4 requirements for DNA polymerase to work?

Answer 12

• All 4 dNTPs as precursors
• DNA template
• A primer with a free 3’ hydroxyl group
• Mg²⁺ cofactor

Question 13

Why does DNA polymerase require a primer with a free 3’ hydroxyl group?

Answer 13

DNA polymerase can only add a dNTP to the 3’ end of a pre-existing or RNA molecule

Question 14

What type of enzyme is primase?

Answer 14

A specialised RNA polymerase

Question 15

What is the function of primase?

Answer 15

Makes short RNA primers

Question 16

Does primase have proofreading?

Answer 16

No

Question 17

What are the requirements for the catalytic site of DNA polymerase III?

Answer 17

• Aspartates
• Mg²⁺
• Triphosphate

Question 18

What is the purpose of single stranded binding proteins?

Answer 18

Used to prevent re-annealing of single stranded DNA so DNA polymerase can move along the strand

Question 19

Which enzyme unwinds double stranded DNA to produce single stranded DNA?

Answer 19

DNA helicase

Question 20

Which direction does DNA polymerase III synthesise DNA?

Answer 20

5’ to 3’

Question 21

Why can DNA polymerase only add nucleotides onto an existing 3’ OH group?

Answer 21

The OH group acts as a nucleophile and attacks the dNTP, breaking the bond between phosphate 2 and 3

Question 22

What is the role of Mg²⁺ in DNA polymerase III?

Answer 22

• Ensures the phosphates on dNTP are positioned correctly in the enzyme
• Activates the OH to make it a better nucleophile

Question 23

Why are aspartates required in the catalytic site of DNA polymerase III?

Answer 23

Put Mg²⁺ in the right direction

Question 24

What is the general function of an exonuclease?

Answer 24

Cleaves nucleotides one at a time from ends of polynucleotides

Question 25

What is the general function of an endonuclease?

Answer 25

Cleaves nucleotides within a polypeptide

Question 26

Describe how DNA is replicated discontinuously (on the lagging strand)

Answer 26

• As DNA is unwound primase makes new primers
• DNA polymerase III produces an okazaki fragment
• DNA polymerase III is replaced by DNA polymerase I which removes the RNA primers and replaces them with DNA
• 3’-5’ proofreading occurs
• DNA ligase catalyses the phosphate linkage

Question 27

What is the general function of a nuclease?

Answer 27

Breaks phosphodiester bonds

Question 28

How are double stranded DNA breaks repaired?

Answer 28

Homologous recombination

Question 29

How does DNA replication get terminated?

Answer 29

• At the termination site an enzyme (called Tus in E.coli) binds to Ter sequences
• The Tus enzyme acts as a physical block and prevents DNA polymerase from moving further