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BIOL 3421 - molecular biochem > repair and recombination > Flashcards

repair and recombination Flashcards

1
Q

define mutation

A

permanent change in DNA sequence

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2
Q

T or F: most mutations are neutral or harmful

A

true

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3
Q

T or F: many mutations are advantageous

A

false; they’re rarely advantageous

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4
Q

list 2 ways in which mutations can be caused by

A

spontaneous mistakes in DNA replication or by environmental mutagens

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5
Q

what is a silent mutation

A

a mutation within non-coding DNA or a mutation that doesn’t affect the resulting protein product

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6
Q

what is the purpose of the ames test

A

to identify chemicals that promote mutations

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7
Q

define carcinogen

A

a chemical that promotes mutations

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8
Q

describe how the ames test works

A

use a bacteria that has been mutated to not produce His. Grow on a His free medium + put the mutagen in the middle. if the mutagen enhances mutations, the bacteria will be mutated to produce His, and we will see growth

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9
Q

T or F: different bacterial strains can be sensitive to different types of mutations

A

true

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10
Q

describe how rat liver extract can be used to predict if the mutagen would harm mammalian DNA as well as bacterial DNA

A

isolate extract that is His-. Plate on medium lacking His and incubate. Pos result = high number of His- to His+ suggests presence of a strong mutagen

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11
Q

what is mismatch repair

A

corrects non-complimentary mismatches made by DNA pol III that were missed by pol III proofreading

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12
Q

what is base-excision repair

A

removes bases with structural abnormalities

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13
Q

what is nucleotide-excision repair

A

removes the entire nucleotide when there is an abnormal base that causes distortions in the overall helical structure

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14
Q

what is direct repair

A

fixes damage or abnormal bases without removing the nucleotide first

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15
Q

dam methylation is used in which type of repair

A

mismatched repair

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16
Q

describe the activities of dam methylation

A

it permits discrimination of the template strand. It methylates both strands, but the template is methylated first so there is a short period in which it’s the only one that’s methylated. This is how we know which base pair in the mismatch was the correct one

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17
Q

during mismatch repair, what happens if both strands had become methylated

A

no repair occurs

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18
Q

during mismatch repair, what happens if neither strand had become methylated

A

repair occurs indiscriminately on broth strands

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19
Q

mismatch repair: which protein complex bind mismatched base pairs

A

MutL-MutS

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20
Q

structure of MutL-MutS?

A

clamp-like

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21
Q

T or F: MutL-MutS binds to mismatched base pairs in an ATP-dependent process

A

true

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22
Q

describe the mechanism of MutL-MutS

A

they bind to the mismatch and then slide along the strand until they reach a hemimethylated site

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23
Q

how far away does the mismatch need to be from the hemimethylated site

A

1000bp

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24
Q

which sequence is methylated by dam methylase

A

5’-GATC-3’

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25
Q

in the 5’-GATC-3’ sequences, which base is methylated + where on that base

A

adenine is methylated on the N6 position

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26
Q

what happens when MutL-MutS reaches the hemimethylated site

A

they encounter MutH

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27
Q

what does MutH do once MutL-MutS arrives at the hemimethylated site

A

MutH has endonuclease activity and will cleave the unmethylated strand on the 5’ side of the G in the GATC sequence

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28
Q

mismatch repair: describe what happens when the mismatch is on the 5’ side of the cleavage site

A

unmethylated strand in unwound and degraded 3’-5’ from nick to mismatch

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29
Q

mismatch repair: describe what happens when the mismatch is on the 3’ side of the cleavage site

A

unmethylated strand in unwound and degraded 5’-3’ from nick to mismatch

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30
Q

for mismatch repair, list 5 enzymes/proteins that are required

A

helicase, exonuclease, pol III, SSBs, and ligase

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31
Q

T or F: mismatch repair is very costly

A

true

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32
Q

why is mismatch repair costly

A

up to 1000 additional dNTPs are required to fix just one mismatched pair

33
Q

in base-excision repair, what type of reaction causes the mistake

A

deamination

34
Q

what is a common deamination

A

C to U

35
Q

T or F: deamination is spontaneous + non-enzymatic

A

true

36
Q

base-excision repair: which enzyme family recognizes base lesions

A

DNA glycosylases

37
Q

what do DNA glycosylases do

A

they recognize base lesions and remove the offending base

38
Q

how do DNA glycosylases remove the offending base in base-excision repair

A

they cleave the N-glycosyl bond

39
Q

cleavage of the N-glycosyl bond leads to which type of nucleotide

A

abasic (lacking a base)

40
Q

list the two types of abasic nucleotides

A

apurinic or apyrimidinic

41
Q

in base-excision repair, how is it fixed if there is no glycosylase

A

the N-b-glycosyl bond between the pentose and the base can hydrolyze

42
Q

base-excision repair: once an abasic nucleotide is present, what happens

A

AP (abasic) endonuclease nicks the DNA backbone either directly 5’ or directly 3’ to the missing base

43
Q

base-excision repair: once an AP endonuclease nicks the DNA backbone beside the missing base, what happens

A

pol I uses its 5’-3’ exonuclease activity to remove a few bases starting from the nick + it replaces them. ligase then seals the nick

44
Q

nucleotide-excision repair: what can cause large distortions in helical structure

A

UV light

45
Q

nucleotide-excision repair: give an example of a large distortion to helical structure that can be caused by UV light

A

thymine dimerization

46
Q

nucleotide-excision repair: describe how much DNA must be removed to make the repair

A

larger chunks need to be removed

47
Q

nucleotide-excision repair: which enzyme is used to make the large repair

A

excinuclease

48
Q

nucleotide-excision repair: describe the actions of excinuclease

A

hydrolyzes two phosphodiester bonds. One nick is created on either side of the lesion

49
Q

nucleotide-excision repair: how many phosphodiester bonds does excinuclease hydrolyze

A

2

50
Q

nucleotide-excision repair: once two nicks are made by excinuclease activity, what happens

A

helicase removes the offending DNA fragment, then pol I replaces them, and ligase seals the nick

51
Q

which disease is caused by defects in nucleotide-excision repair machinery

A

Xeroderma Pigmentosum

52
Q

describe the symptoms of Xerodoma Pigmentosum

A

extreme sunlight sensitivity, freckling, increased risk of skin cancers, irritated eyes/impaired vision, increased risk of eye cancers

53
Q

name the enzyme that is very good at repairing thymine dimers

A

photolyase

54
Q

which organisms have photolyase

A

bacteria and plants

55
Q

T or F: humans synthesize photolyase

A

false! only plants and bacteria do

56
Q

when is direct repair used (ie what was the damaged caused by)

A

direct repair is used to repair damage caused by alkylation

57
Q

direct repair: give an example of alkylation that occurs

A

guanine –> O6-methylguanune

58
Q

direct repair: what does O6-methylguanine base pair with

A

pairs with T instead of C

59
Q

direct repair: describe the mechanism

A

methyltransferase can transfer the methyl of O6-methylguanine to one of it’s cysteines. This inactivates the protein permanently but regenerates the guanine

60
Q

when do double stranded breaks and ssDNA occur

A

occur when a replication fork runs into already damaged DNA

61
Q

what happens when the replication fork runs into already damaged DNA

A

the replisome usually falls off

62
Q

how do double stranded breaks get repaired

A

by a mechanism involving homologous recombination

63
Q

describe how homologous recombination is involved with fixing DNA with no template

A

the accurate nucleotides are determined from the homologous chromosome

64
Q

DNA damage with no template: which enzyme is used to replicate DNA with lesions

A

DNA pol V

65
Q

describe the error rate of DNA pol V

A

it can replicate over lesions that would normally stall other polymerases, but with a much higher error rate

66
Q

what is homologous recombination

A

rearrangement of genetic info between chromosomes that share extended regions of (almost) identical sequences

67
Q

homologous recombination: describe the events that occur after a replication fork collapses upon encountering a DNA nick and the 5’ end of the dsDNA is degraded

A

the exposed 3’ end is bound to recombinase, which facilitates another 3’-5’ strand to invade and undergo complimentary base pairing with the now exposed 3’ end. The structure can migrate along the strand and create an X-like crossover known as a Holliday intermediate. Nucleases cleave the Holliday, and the strands are re-ligated, and the replication fork is re-established

68
Q

homologous recombination: what protein is responsible for the initial 5’ degradation

A

RecBCD

69
Q

homologous recombination: which types of activity does RecBCD have

A

both helicase and exonuclease

70
Q

homologous recombination: which is the recombinase that promotes strand invasion

A

RecA

71
Q

homologous recombination: what does RecA do

A

it forms a long filament on ssDNA with up to several thousand subunits

72
Q

homologous recombination: what does RecBCD do

A

assists in removing the SSBs from ssDNA and then allowing RecA to bind

73
Q

list the 3 purposed of homologous recombination

A

helps repair DNA damage, helps to physically link chromatids for proper division during meiosis I, enhances genetic diversity

74
Q

what does meiosis I separate

A

homologous chromosomes

75
Q

T or F: cohesins keep homologues together

A

false! there are no cohesins. only sister chromatids have cohesins

76
Q

what is aneuploidy

A

incorrect number of chromosomes in a daughter cell

77
Q

in humans, when does egg production begin

A

week 12 of gestation (before female is even born)

78
Q

describe meiosis from gestation to ovulation

A

meiosis is initiated in the fetal germ line cells. Cells proceed through most of meiosis I, where chromosomes line up and generate cross overs. The process then stops with the crossovers in place, until just before ovulation

79
Q

T or F: the longer the arrested stage of meiosis, the higher risk of aneuploidy when the homologues finally separate

A

true

BIOL 3421 - molecular biochem (20 decks)

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