DNA Polymerases in DNA repair

Question 1

List the five enzymes involved in DNA rreplication.

Answer 1

DNA helicase

DNA primase

DNA polymerase

DNA ligase

Nuclease

Question 2

What does nuclease do?

Answer 2

Removes and incorrect nucleotides

Question 3

What are the three main polymerases used in prokaryotic DNA replication?

Answer 3

DNA pol I

DNA pol II

DNA pol III

Question 4

What does DNA pol II primarily do?

Answer 4

Repair

Question 5

What does DNA pol III primarily do?

Answer 5

DNA synthesis

Question 6

What does DNA pol I do in the forward direction?

2

Answer 6

Exonuclease activity

Nick translation (random nicks in DNA)

Question 7

Define DNA damage.

Answer 7

Any alteration to DNA that distorts its geometry

Question 8

What causes DNA damage?

2

Answer 8

Endogenous agents

Exogenous agents

Question 9

Give two examples of endogenous damage.

Answer 9

Replication errors

Metabolism

Question 10

Give three examples of exogenous damage.

Answer 10

Toxins

Chemicals

UV radiation

Question 11

Characterise DNA damage.

Answer 11

Single strand damage (SSD)

Double strand damage (DSD)

Question 12

What does the type of DNA damage depend on?

Answer 12

Depends on the type of agent and the severity of the agent that caused the damage

Question 13

What causes SSD damage?

3

Answer 13

Endogenous replication errors

Oxidative chemicals

Non-ionising radiation e.g. UV

Question 14

What causes DSD damage?

2

Answer 14

Antitumor agents e.g. Cisplatin

Ionising radiation e.g. Gamma rays

Question 15

What are the five types of single strand damage?

Answer 15

Base Mismatch

Depurination

Deamination

Thymine dimers

Bulky adducts

Question 16

What is base mismatch SSD?

Answer 16

Distorted geometry of DNA (associated with errors in replication)

Question 17

What is depurination SSD?

Answer 17

Reaction resulting in the removal of Purines A or G

Question 18

What is deamination SSD?

Answer 18

Loss of an amino group from Cytosine to produce uracil or loss from Adenine to produce hypoxanthine

Question 19

What is formed when cytosine loses an amino acid?

Answer 19

Uracil

Question 20

What is formed when adenine loses an amino acid?

Answer 20

Hypoxanthine

Question 21

What is thymine dimers SSD?

Answer 21

Covalent linkage of 2 adjacent thymines together (associated with UV radiation)

Question 22

What is bulky adducts SSD?

2

Answer 22

Occur due to DNA intercalalting agents such as anticancer agent Cisplatin

Cisplatin binds to bases and forms bulky lesions in the DNA helix

Question 23

What is a mutation?

Answer 23

Randomly produced permanent change in a sequence of DNA

Question 24

When can a single permanent mutation severely compromise an organism’s fitness?

Answer 24

If the change occurs in a vital position in the DNA sequence

Question 25

What happens if a mutation occurs in a protein coding portion of DNA?

Answer 25

A protein with an altered amino acid sequence will result -> defective protein

Question 26

Give an example of a disease caused by a mutation of the DNA resulting in incorrect amino acid sequence of a protein.

Answer 26

Sickle cell anaemia

Question 27

What are the three repair processes for SSD?

Answer 27

Mismatch repair (MMR) for repairing Base excision repair (BER) Nucleotide excision repair (NER)

Question 28

What is MMR?

Answer 28

Repairs rare mistakes made during DNA replication process (endogenous) that escape DNA polymerase III and I proofreading

Question 29

What is BER?

Answer 29

Repair of modified DNA bases from depurination or deamination

Question 30

What is NER?

Answer 30

Repair of bulky lesions and distortions in DNA

Question 31

Explain how MMR works. | 4

Answer 31

Replication machines makes a copying mistake Mismatch base pair occurs Mismatch repair proteins recognise the mismatch and remove the new strand Missing strand is now resynthesized

Question 32

How does MMR repair a nick? | 2

Answer 32

DNA mismatch repair proteins recognise a nick in DNA DNA pol I repairs the nick and replaces the DNA at the cleaved site

Question 33

How does Ber-Base Excision repair work? | 5

Answer 33

A DNA glycosylase recognises a damaged base and cleaves between the base from the deoxyribose in the backbone leaving an AP site An AP endonuclease cleaves the phosphodiester backbone near AP site creating a nick in the strand Phosphodiesterase removes the AP sugar and phosphate DNA Pol I replaces the damaged bases The phosphodiester nick is sealed with DNA ligase

Question 34

What does DNA glycosylase do? | 2

Answer 34

Recognises a damaged base Cleaves between the base from the deoxyribose in the backbone leaving an AP site

Question 35

What does AP endonuclease do?

Answer 35

Cleaves the phosphodiester backbone near AP site creating a | nick in the strand

Question 36

What does Phosphodiesterase do?

Answer 36

Removes the AP sugar and phosphate

Question 37

How does Ner-Nucleotide Excision Repair work?

Answer 37

Damaged DNA recognised and removed by one of a variety of different nucleases Nucleases first scan and cleave the phosphodiester bonds that join damaged nt to rest of DNA molecule. Results in small gap on 1 strand of DNA double helix Repair DNA polymerase II binds to 3’ OH end of cut DNA strand and fills the gap Nick remains in sugar-phosphate backbone until it is sealed by DNA ligase

Question 38

Give an example of a nuclease

Answer 38

The UvrABC endonuclease enzyme complex consisting of four Uvr proteins: UvrA, UvrB, UvrC, and UvrD (DNA helicase II))

Question 39

What is UvrABC?

Answer 39

An endonuclease enzyme complex consisting of four Uvr proteins: UvrA, UvrB, UvrC, and UvrD (DNA helicase II))

Question 40

What does UvrABC do?

Answer 40

Scan and cleave the phosphodiester bonds that join damaged nt to rest of DNA molecule.

Question 41

What does the cleavage of a phosphodiester bond do?

Answer 41

Results in small gap on 1 strand of DNA double helix

Question 42

What fills the gap caused by UvrABC?

Answer 42

Repair DNA Polymerase II

Question 43

What is the final step in repair of a nick?

Answer 43

DNA ligase seals the nick