Lecture 16: Replication and Repair

Question 1

Human genome replication time

Answer 1

-8 hours
-200,000 bp per second
-multiple replication origins

Question 2

Error rate of replication

Answer 2

one error per billion bps

Question 3

DNA polymerases

Answer 3

-synthesize DNA
-add nucleotide
-conserve genetic info
-requires 3’-OH of primer
-delta

Question 4

DNA synthesis direction

Answer 4

occurs in 5 to 3 while moving 3 to 5

Question 5

dNTPs

Answer 5

-deoxyribonucleotides
-substrates for DNA synthesis
-not enough = replication STOPS

Question 6

Phosphodiester bond

Answer 6

-3’OH group with a-phosphate of dNTP
-releases a pyrophosphate (ppi)

Question 7

Leading strand

Answer 7

-new template sequence in front of the synthesis
-continuous

Question 8

Lagging Strand

Answer 8

-new template behind the synthesis
-okazaki fragmentsl

Question 9

ligase

Answer 9

link okazaki frgments

Question 10

Unwinding

Answer 10

-helicase unwinds DNA
-exposed DNA stabilized by single-strand binding (SSB) proteins

Question 11

Priming

Answer 11

-primase synthesizes short RNA primer

Question 12

Strand elongation

Answer 12

DNA polymerases synthesize DNA

Question 13

Primer removal

Answer 13

RNA primers digested by RNase H

Question 14

Gap Filling

Answer 14

DNA polymerases synthesize DNA to end of okazaki fragment in front

Question 15

Ligation

Answer 15

ligase seals nicks in DNA strand

Question 16

Proofreading

Answer 16

-DNA polymerases has 3 to 5 exonuclease activity
-removes newly added nucleotide
-proofreads nucleotide at 3’ end

Question 17

proper nucleotide at 3’ end

Answer 17

suppresses proofreading

Question 18

mismatched nucleotide at 3’ end

Answer 18

supresses synthesis

Question 19

processivity

Answer 19

-DNA polymerases may dissociate from DNA before completion of synthesis
-rebinding of DNA polymerase slows replication

Question 20

enhanced processivity

Answer 20

increased rate of DNA synthesisS

Question 21

Sliding Clamp

Answer 21

-PCNA (proliferating cell nuclear antigen) holds down DNA polymerase
-suppresses dissociation
-increases processivity

Question 22

Replisome

Answer 22

complex of polymerases working on the leading and lagging strands

Question 23

Ending replication problem

Answer 23

-DNA linear
-5’ end of lagging strand cannot be replicated bc no primer can be made
-DNA shortened by 50-100 bp per replication

Question 24

telomere

Answer 24

-located at end of chromosomal DNAs
-repeat TTAGGG w/o genetic info
-lost after 40-60 replications = cell death

Question 25

Hayflick limit

Answer 25

-40-60 replications before telomere is lost =cell death

Question 26

Telomerase

Answer 26

-ribonucleoprotein complex with RNA -adds TTAGGG to 3' end of telomere using RNA sequence as template -enlongates telomere -increases lifespan of cells

Question 27

Cancer cell immortality

Answer 27

-reactivation of telomerase

Question 28

Polymerase chain reaction (PCR)

Answer 28

-amplifies DNA fragment tagged by DNA primers in vitro -3 step cycle repeats 25-30 times -cloning and detection (PCR test for covid)

Question 29

PCR phases

Answer 29

1. denaturing (94degrees) 2. Anneling (50) 3. extension. (72) -each at different temp -requires thermocycler and a heat-resistant polymerase

Question 30

DNA damage causes

Answer 30

-reactive oxygen -deamidation -depurination

Question 31

reactive oxygen species

Answer 31

-hydroxy radical (HO-) -superoxide (O2-)

Question 32

Depurination

Answer 32

-several thousands of purine lost a day -generates abasic (AP) sites -missing bp

Question 33

Deamidation

Answer 33

N to =O

Question 34

External causes of DNA damage

Answer 34

-carcinogens -UV radiationca

Question 35

carcinogens

Answer 35

-mustard gas -N-nitroso in meat -chemo -benzopyrene

Question 36

UV RADIATION

Answer 36

-pyrimadine dimers

Question 37

DNA repair

Answer 37

-base-excision -nucleotide-excision -mismatch -demethylation -photoreactivation -double-strand breakssdessdessdeesdesdesdewsdesxcfdscgresxcvgrew

Question 38

base-excision repair

Answer 38

-damage base removed -forms AP site -AP site removed -gap resynthesized

Question 39

nucleotide-excision repair

Answer 39

-stretch of nucleotides with damage is removed -gap resynthesized

Question 40

Xeroderma Pigmentosum (XP)

Answer 40

-autosomal recessive genetic disease -defect in nucleotide-excision repair -photosensitive and susceptive to skin cancer

Question 41

mismatch repair

Answer 41

-fix mismatch pair -similar to nucleotide-excision -mutation occurs -methylation may guide identification of parental strand

Question 42

direct demethylation

Answer 42

removes methyl group from O-methylguanine

Question 43

Photoreactivation

Answer 43

reverts pyrimidine dimer

Question 44

Double-strand break repair

Answer 44

-10 breaks per cycle -homologous recombination (Accurate) -non-homologous end-joining (error-prone)

Question 45

p53

Answer 45

-control DNA repair -transcription factor responding to damages

Question 45

p53 functions

Answer 45

-activates DNA repair proteins -arrests cell cycle -may initiate programmed cell death if too much damage

Question 45

p53 tumor suppressor

Answer 45

-failure of programmed cell death may cause cancer -frequently mutated in human cancer