mechanism of DNA replication

practice questions

LECTURE 13: DNA: STRUCTURE & REPLICATION II

MECHANISM OF DNA REPLICATION

DNA replication requirements... 1. H bonds between bases must be broken 2. chain separation/unwinding 3. available pools of 4 dNTPs: A = T, C G 4. enzymes

E. coli... ~4.7 × 107 bp (47,000,000) molecule has ~ 4.7 × 106 turns (10 bp/turn) 1500 bp/s, ~ 20 min to divide with 1 origin

MECHANISM OF DNA REPLICATION DNA polymerase I (pol I)... several types of activity 1. polymerase catalyzes chain growth 5' 3' 2. exonuclease removes mismatches 3' 5' 3. exonuclease degrades double stranded DNA 3' 5'

MECHANISM OF DNA REPLICATION DNA polymerase III (pol III) ... holoenzyme of 20+ subunits 1. polymerase catalyzes chain growth 5' 3'

MECHANISM OF DNA REPLICATION prokaryotes have a single origin of replication bidirectional replication with 2 forks single terminus ~ 20 minutes (*... book says 40)

MECHANISM OF DNA REPLICATION E. coli origin of replication tandem repeat consensus sequence protein binding sites

MECHANISM OF DNA REPLICATION

replication to completion...

MECHANISM OF DNA REPLICATION eukaryotes have multiple origins of replication bidirectional replication with 2 forks 1.4 h (yeast) to 24 h (cultured animal cells)... or more issues...

linear vs circular coordination of multiple chromosomes

pulse/chase and autoradiography ...

MECHANISM OF DNA REPLICATION eukaryotes have multiple origins of replication bidirectional replication with 2 forks 1.4 h (yeast) to 24 h (cultured animal cells)... or more issues...

linear vs circular coordination of multiple chromosomes

pulse/chase and autoradiography ... Drosophila

MECHANISM OF DNA REPLICATION DNA structure replication

enzymes functions

MECHANISM OF DNA REPLICATION DNA structure

replication

5' 3'

enzymes functions

3'

5'

3'

5'

MECHANISM OF DNA REPLICATION DNA structure

replication

5' 3'

phosphodiester bond

enzymes functions

MECHANISM OF DNA REPLICATION helicases disrupt H-bonds binding strands of

DNA double helix

MECHANISM OF DNA REPLICATION single-stranded binding proteins (SSBs) stabilize

DNA and prevent formation of duplex

MECHANISM OF DNA REPLICATION topoisomerases relax supercoiling of DNA as

it unwinds... e.g., DNA gyrase

MECHANISM OF DNA REPLICATION topoisomerases relax supercoiling of DNA as

it unwinds... e.g., DNA gyrase

MECHANISM OF DNA REPLICATION DNA gyrase -

catalyzed supercoiling

MECHANISM OF DNA REPLICATION DNA polymerase catalyzes replication 5' 3'

MECHANISM OF DNA REPLICATION DNA polymerase catalyzes replication 5' 3' extends but cannot initiate replication ...

MECHANISM OF DNA REPLICATION RNA primase enzyme

MECHANISM OF DNA REPLICATION priming synthesis

primase enzyme

synth. short (~30 bp) RNA primer

primase + DNA + proteins = primosome

enzymes functions

MECHANISM OF DNA REPLICATION primase + DNA + proteins = primosome

MECHANISM OF DNA REPLICATION leading & lagging strand

replication is 5' 3'

replication moves along template 3' 5'

DNA pol III most synthesis

DNA pol I fills gaps on lagging strand

DNA ligase seals gaps

MECHANISM OF DNA REPLICATION replication fork

lagging strand

a) RNA primer growth

b) DNA elongation as Okazaki fragments

c) RNA primer removed by DNA pol I

d) DNA ligation

MECHANISM OF DNA REPLICATION dimeric DNA pol III holoenzyme model

MECHANISM OF DNA REPLICATION

chromosome ends

leading strand has no problem...

lagging strand cannot add RNA primer at end... leaving gaps

telomeres = DNA repeats made by...

telomerase

MECHANISM OF DNA REPLICATION telomerase has RNA template for making telomeres

telomere-specific DNAprobe (yellow)

MECHANISM OF DNA REPLICATION enzyme summary...

MECHANISM OF DNA REPLICATION

you should know names and functions of:

topoisomerase (e.g., DNA gyrase) single-stranded binding proteins (SSBs) DNA helicase DNA pol I DNA pol III DNA pol III holoenzyme RNA primer RNA primase

primosome Okazaki fragment DNA ligase telomerase

PRACTICE QUESTIONS

you should be able to do

CH7, p. 250 # 1 - 25