DNA repair

Question 1

sites of potential damage to DNA

Answer 1

Oxidative damage
Spontaneous hydrolysis
Nonenzymatic methylation by SAM

Question 2

how to fix ds break

Answer 2

Non homologous end joining: doesn’t require a repair template

Question 3

how to Enhance replication fidelity

Answer 3

mismatch repair: occurs only in coordination with DNA replication

Question 4

how to repair of spontaneous base hydrolysis, chemical damage, or other forms of damage

Answer 4

Direct reversal by photolyase (UV-light)
Excision repair (base or nucleotide excision repair)
Base is smaller repair, just one
nucleotide excision is larger

Question 5

ds break repair details

Answer 5

NHEJ
Ku70/80 binds ends, recruits proteins that brings ends together, DNA ligase seals gaps
This must happen quickly before nucleases notice break and degrade ends
This repair can occur during anytime in the cell cycle

Question 6

mismatch repair details

Answer 6

new strand has error

1) MutS recognizes error, binds recruits MutL
2) MutH binds daughter strand, mismatch is looped out, MutH nicks daughter strand
3) MutL recruits UvrD (helicase: unwinds 3’ to 5’)
4) exonuclease digests ssDNA tail
5) DNA pol III + SSBP fill in correct nucleotide
6) DNA ligase seals gap and daughter strand is methylated

Question 7

what does mismatch repair require

Answer 7

hemi-methylated DNA helix

Question 8

UV irradiation

Answer 8

can cause dimerization of pyrimidines → this distorts DNA

Pyrimidine dimers can be repaired by direct reversal by photolyases or by excision repair

Question 9

base excision repair

Answer 9

Excises a single base (1-3 or 4)
DNA glycosylase removes the base.
AP (apurinic site) endonuclease cleaves the backbone and an exonuclease removes the deoxy-nucleotide
DNA pol I fills the gap and ligase seals the nick
This may be used to repair UV-light induced damage, spontaneous hydrolysis/deamination, or low level alkylating agent induced damage

Question 10

nucleotide excision repair

Answer 10

Excises a larger region of DNA
This may be used to repair UV-light induced damage, major damage by alkylating agents, or other major sites of RNA damage

Question 11

uracil in DNA

Answer 11

highly mutagenic

Question 12

spontaneous modifications

Answer 12

deamination of cytosine to uracil

If uracil were a normal base in DNA, this would cause problems for repair enzymes

Question 13

how do we recognize uracil in DNA

Answer 13

For example, repair enzymes would be unable to discern if the correct pair (from a G-U mismatch) should be G-C or A-U
BUT, bc uracil is not a normal DNA base (and thymine is), any uracil present in DNA comes from deaminated cytosine

Question 14

how do we fix uracil in DNA

Answer 14

Uracil-DNA glycosylase is responsible for excising the base, which is correct through base excision repair. This also helps remove uracil that is incorporated into DNA via the presence of dUTP

Question 15

point mutations

Answer 15

substitution of one base pair for another

May be caused by errors in DNA replication (rare) or by chemicals that alter the structure of a base

Question 16

insertions and deletions

Answer 16

(indels) may be caused by errors in replication (Replication slipping) or by the formation/incorrect repair of double-stranded breaks

Question 17

point mutations due to base mispairing

Answer 17

transitions and transversions

Question 18

transitions

Answer 18

a purine (or pyrimidine) is replaced by a different purine (or pyrimidine)

Question 19

transversions

Answer 19

a purine is replaced by a pyrimidine (or vice versa)

Question 20

how do point mutations arise

Answer 20

rare tautomerization, the presence of base analogs, spontaneous hydrolysis, exposure to chemical mutagens that modify base structure

Question 21

rare tautomerization

Answer 21

A-imino pairs with cytosine instead of thymine

Leads to AT to GC transition in the next generation

Question 22

base analogs can induce point mutations

Answer 22

5-bromouracil (5-BU) is a thymine analog; pairs with guanine
This causes an A to G transition
Enol form of 5-BU can be incorporated in place of cytosine. If it returns to its keto form (T-like), this causes a mispairing from CG to TA

Question 23

2-AP

Answer 23

2-AP is an adenine analog that pairs with thymine or cytosine
This can cause a T to C transition

Question 24

hypoxanthine

Answer 24

Arises from spontaneous deamination of adenine, and it pairs with cytosine
This causes a TC transition

Question 25

chemical mutagens

Answer 25

Chemical mutagens modify bases, changing their base-pairing qualities
Nitrous acids causes oxidative damage
Nitrous acid deaminates cytosine and adenine

Question 26

nitrosamines modify bases in 2 ways

Answer 26

Nitrosamines react to form nitrous acid

Nitrosamines can alter cytosine so that it pairs with adenine

Question 27

hydroxylamine

Answer 27

modifies cytosines

Question 28

alkylating agents

Answer 28

modify bases by adding alkyl-groups (methyl or ethyl groups) to different positions on bases

Question 29

insetions and deletions

Answer 29

Addition or removal of one or more base pairs
If in the coding sequence, these can lead to frameshift mutations that alter the amino acid sequence or lead to a premature stop codon

Question 30

intercalating agents

Answer 30

such as ethidium bromide or acridine orange, bind to DNA

These cause distortions that alter the distance between base pairs

Question 31

radiation damage

Answer 31

can also induce double stranded breaks; indels occur if this damage is not repaired precisely

Question 32

which is worse: DNA or RNA errors?

Answer 32

DNA bc *We make a lot more RNA, so there is more tolerance for errors
One error in one protein is fine if we make a ton of proteins