DNA replication and repair

Question 1

nucleoside analogs

Answer 1

nucleosides that lack the 3’-OH group and nucleosides with modified sugars (arabinose instead of ribose) both act as chain terminators in DNA replication. They are easily incorporated but impossible to extend due to their geometry, thus, they can be used as anticancer and antiviral drugs to target DNA polymerase and prevent DNA pol from elongating the chain.

Question 2

topoisomerase type 1

Answer 2

creates single stranded breaks in DNA, drugs that inhibit topoisomerase 1 cause ss breaks in DNA.

Question 3

topoisomerase type 2

Answer 3

also known as gyrase in prokaryotes, creates double stranded breaks in DNA; drugs that inhibit this inhibit negative supercoiling in bacteria = death, or stabilize broken DNA after the cut (anticancer)

Question 4

telomerase

Answer 4

enzyme that elongates telomeres (hexameric repeats at both ends of dsDNA, longer on 3’ end)
**increased telomerase activity as seen in stem cells and cancer cells allows for immortality, thus telomerase is a target for cancer therapy

**somatic cells do not have telomerase and thus can only divide 20-60 times before death

Question 5

mismatches

Answer 5

caused by mistakes made by DNA polymerase during replication; occur more often in microsatellite regions due to the di- and tri-nucleotide tandem repeats that create stable loop formations.

**repaired with MMR, defective MMR = Lynch syndrome

Question 6

spontaneous non-enzymatic alterations

Answer 6

includes deamination of bases, loss of bases.

*repaired with BER

Question 7

deamination of bases

Answer 7

usually cytosine to uracil, or methylated cytosine to thymine. Methylated cytosines are hotspots for point mutations due to this conversion.

*repaired with BER

Question 8

UV radiation

Answer 8

causes cross linking and dimerization of adjacent thymine residues, which causes distortion of the backbone.

*repaired with NER, defect in NER = xeroderma pigmentosum

Question 9

ionizing radiation

Answer 9

gamma rays, x-rays; causes ds and ss breaks in DNA, repaired with NHEJ and HR

Question 10

Ame’s test for mutagenicity

Answer 10

test to determine a chemical’s mutagenicity, uses mutated Salmonella that cannot produce histidine in a hisitidine-free media growth plate - if salmonella grows, the chemical has created mutations

Question 11

alkylation

Answer 11

usually due to chemical alkylating agents that add an alkyl group to guanine

Question 12

direct reversal

Answer 12

repair mechanism that removes alkyl groups from guanine using MGMT protein

Question 13

BER

Answer 13

recognizes base deamination and oxidative damage; glycosylase removes base, then APE makes initial cut, then AP lyase removes base, then DNA pol and ligase reseal.

Question 14

MGMT

Answer 14

protein that removes alkyl group from guanine in direct reversal repair

Question 15

MMR

Answer 15

repairs base-base mismatch and small indels caused by mistakes by DNA pol during replication. (Thus important for microsatellite stability!)

• Enzymes recognize daughter strand because it is non-methylated.
• Uses helicase.

Defective MMR = Lynch syndrome =cancer.

Question 16

NER

Answer 16

activated by helix distortion, i.e. when UV radiation causes thymine dimerization.
**2 pathways, global protein-NER and transcription-coupled NER.

**Defects cause xeroderma pigmentosum.

Question 17

homologous recombination (HR)

Answer 17

primary pathway for DSB repair in prokaryotes, also used in eukaryotes. Less error prone than NHEJ. Occurs after replication and uses the DNA sister chromatid as a template.

**normally promoted by BRCA1+2

Question 18

non-homologous end joining (NHEJ)

Answer 18

primary pathway for DSB repair in eukaryotes, more error prone but does not use sister chromatid and occurs in G1 phase.

**if BRCA1+2 mutations are present, HR will be blocked and NHEJ pathway takes over, resulting in more errors - leads to cancer

Question 19

Lynch syndrome

Answer 19

defect in MMR genes, increased risk for colorectal and endometrial cancer

Question 20

xeroderma pigmentosum

Answer 20

defect in NER genes, causes UV intolerance and 2000x increase risk of fatal skin cancer

Question 21

BRCA1+2

Answer 21

normally promote HR, but if mutated HR will not proceed and NHEJ takes over = increased errors and increased risk for cancer

Question 22

defects in NHEJ

Answer 22

causes SCID because unable to do VDJ recombination and produce antibodies