DNA Repair

Question 1

different categories of DNA damage?

Answer 1

replication errors
spontaneous damage
environmental damage

Question 2

what do mutations lead to?

Answer 2

cancer & heritable disease
restored DNA
senescence/apoptosis

Question 3

how is DNA damage fixed?

Answer 3

direct reversal
base excision repair
nucleotide excision repair
mismatch repair

Question 4

how many mismatches per cell division?

Answer 4

100 mismatches per cell division

Question 5

how many lesions per cell per day?

Answer 5

10^5

Question 6

lesion definition?

Answer 6

modification to DNA

Question 7

mutation definition?

Answer 7

misincorporation of nucleotides during replication

Question 8

how does DNA repair affect no of lesions per cell per day?

Answer 8

Reduces 1,000 fold

Question 9

transition mutations?

Answer 9

A  G (purines)
C  T (pyrimidines)

Question 10

transversion mutations?

Answer 10

A C or T

G C or T

Question 11

replication errors?

Answer 11

mismatches
tautomerisation
strand slippage
intercalation

Question 12

spontaneous and environmental damage?

Answer 12

hydrolysis (depurination/deamination)
oxidation 
alkylation 
bulkyl adduct formation
radiation

Question 13

error rate of typical DNA polymerase?

Answer 13

1 in 10^5 nucleotides

1 in 10^7 after DNA repair

Question 14

what are the wobble pairs?

Answer 14

A-C especially in lower pHs, maybe in nucleus

G-T

Question 15

are the wobble pairs isomorphic?

Answer 15

no, they don’t overlay. Allows some enzymes to recognise that they aren’t the correct base pair

Question 16

what is tautomerisation?

Answer 16

alternate hydrogen positioning.

usual keto form of T can be in the alternative enol form, can allow it to pair with G

imino A can also pair with C

Question 17

what can enol T pair with?

Answer 17

G

Question 18

what can imino A pair with?

Answer 18

C

Question 19

what is strand slippage?

Answer 19

newly synthesised strand can slip out slightly and cause an extra nucleotide to be incorporated

or for deletion - template strand slips

Question 20

where is strand slippage prevalent?

Answer 20

in trinucleotide repeats and Huntington’s

Question 21

what is intercalation?

Answer 21

causes insertions.
intercalation is exhibited by flat/aromatic compounds which can sit between base pairs.
disrupt the replication machinery and can lead to insertion mutations (e.g. frameshift)

Question 22

example of intercalation substance?

Answer 22

ethidium bromide found in dyes and cigarette smoke

Question 23

what is an insertion due to transposons?

Answer 23

elements which come from viruses can catalyse their removal and insertion into different regions of genes

Question 24

what is a transposon?

Answer 24

cut and paste

Question 25

what is a retrotransposon?

Answer 25

copy and paste

Question 26

what can chemical modifications lead to?

Answer 26

``` mismatches structural alterations to double helix single/double stranded breaks blockage/stalling of machinery influence gene expression by altering chromatin architecture ```

Question 27

what percentage of genetic mutations are caused by simple replication errors?

Answer 27

66%

Question 28

what is depurination?

Answer 28

spontaneous hydrolysiss of the glycosidic bond between a purine and the formation of an ABASIC site 20 times more likely to happen with purines increased susceptibility to strand breaks

Question 29

what is deamination?

Answer 29

converts C to U by hydrolysis adenine --> hypoxanthine guanine --> xanthine 5-methylcytosine --> T

Question 30

why does DNA contain T, but RNA contains U?

Answer 30

to detect cytosine deamination

Question 31

what is base oxidation by reactive oxygen species?

Answer 31

guanine can often be attacked at 8 position and add oxygen. leads to bond formation with A

Question 32

what is base oxidation by reactive nitrogen species?

Answer 32

adenine --> hypoxanthine which pairs with C

Question 33

what is base alkylation?

Answer 33

transfer of methyl, ethyl or alkyl groups to nitrogen or oxygen of base often used in chemotherapy G pairs with T forms 3-methyl A

Question 34

what does 3-methyl A do?

Answer 34

blocks polymerase from processing

Question 35

what is base adduct formation?

Answer 35

modifies N7 of guanine and causes GC to TA transversions

Question 36

example of base adduct chemical?

Answer 36

alfatoxin B

Question 37

what makes aflatoxin B reactive?

Answer 37

metabolism by cytochrome P450 in liver

Question 38

what is base dimerisation?

Answer 38

caused by UV exposure and causes dimerisation of pyrimidines next to each other Cs and Ts really distort the DNA so easily fixed

Question 39

Ames test?

Answer 39

examines the mutagenic potential of a compound contains defective histidine gene but requires histidine to live mix with mutagen frameshift mutations will reactivate histidine gene and colonies will grow

Question 40

mechanisms for repairing replication errors and DNA damage?

Answer 40

``` direct reversal base excision repair nucleotide excision repair mismatch repair double strand break repair translesion synthesis ```

Question 41

direct reversal?

Answer 41

acts directly on modification (alkylation/UV) human example removes a methyl group using MGMT. Irreversible reaction once methyl is transferred (suicidal enzyme). Methylated form of MGMTis a transcription activator, upregulates synthesis of other repair proteins

Question 42

base excision repair?

Answer 42

removes specific small non helix distorting base lesions. (deamination/oxidation/alkylation) removes base. DNA glycosylase generates an abasic site. AP endonuclease leaves a hydroxyl group. ligase/DNA pol I seal the gap e.g. uracil DNA glycosylase

Question 43

MutM?

Answer 43

removes 8-oxo-G from the GC lesion

Question 44

MutY?

Answer 44

removes A opposite 8-oxo-G following DNA replication

Question 45

MutT?

Answer 45

breaks down 8-oxo-dGMP and ppi before DNA synthesis

Question 46

nucleotide excision repair?

Answer 46

cleaves backbone either side, removes oligonucleotide (bulkyl, helix distorting lesions) endonuclease/helicase then DNA poly I/ligase UvrABCD recognise and remove pyrimidine dimers UvrA recognises lesion UvrB opens and binds DNA UvrC creates incisions either side of lesion UvrD excises the oligonucleotide can see a big difference in the DNA surface when there is a TT dimer

Question 47

difference in nucleotide excision repair in humans?

Answer 47

single stranded segment is longer can get recruited to an RNA polymerase which is stalled at the site of a lesion

Question 48

mismatch repair?

Answer 48

error reducing process of 1000 fold repairs main GT, GG, AC and CC mismatches removes a long stretch of nucleotides MutSLH recognises distortion. straight after replication, daughter strand isn't methylated yet so this means it can find the mismatch and replace the unmethylated DNA MutH nicks the daughter strand, MutS recognises the distortion. MutL is the communicator between the two exonuclease, UvrD (helicase) and DNA polymerase

Question 49

double strand break repair?

Answer 49

before the two sister chromosomes have been generated, NHEJ protects and processes the broken ends and joins them together but usually results in small deletions and is mutagenic

Question 50

translesion polymerases?

Answer 50

non processive, template dependent polymerases that synthesis acrss a blockage, introduce nucleotides independent of base pairing some enzymes fix certain lesions - open active site so allows any nucleotide to be added

Question 51

p53 protein?

Answer 51

transcription factor that regulates the expression of genes involved in growth arrest, DNA repair and apoptosis 50% of cancers have a defective p53

Question 52

Xeroderma Pigmentosum?

Answer 52

Increased sensitivity to UV light | 1000 fold increased risk of skin cancer