LECTURE 10 - DNA replication - overview Semiconservative...

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DNA replication - overview p 530-550 Semiconservative replication (Meselsohn and Stahl)
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DNA replication - replication fork p 530-550 leading strand lagging strand Replication foci
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DNA replication - replication fork, a complex factory p 530-550
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DNA replication - topoisomerase p 530-550 DNA in cells is negatively supercoiled: assists replication process
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DNA replication - topoisomerase p 530-550 Type I topoisomerase : -nick mechanism -covalent enzyme intermediate -no ATP needed Inhibited by irinotecan -an analog of camptothecin (shown) -an anticancer drug -nicked DNA does not reclose -DNA does not replicate -rapidly dividing cells die
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DNA replication - topoisomerase p 530-550 Type II topoisomerase: -strand breaks: passing a strand through the gap -covalent enzyme intermediate -requires ATP +ATP Bacterial topoisomerase II = gyrase -inhibited by ciprofoxacin -causes more DNA cleavage -DNA does not replicate -bacteria die (good against Bacillus anthacis = anthrax) http://www.youtube.com/watch?v=k4FbPUGKurI&Feature=related
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DNA replication - helicase and SSB p 530-550 Helicases: -separate ds DNA into ss DNA -uses ~1ATP/5 base-pairs -35 - 730 base-pairs/second -rotates like F 1 F 0 -ATPase? SSB: -coat single-stranded DNA -prevents reannealing -interacts via Lys (-->phosphate) and Phe/Trp (-->stacking with bases)
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DNA replication - two problems for DNA polymerase p 530-550 1. DNA polymerase can only extend -to initiate, use a short stretch of RNA = primer -put there by primase 2. DNA polymerase must coordinate leading and lagging strand synthesis (5’ --> 3’) -continuous for leading strand -discontinuous for lagging strand (Okazaki fragments) -tight coordination between the two
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DNA replication - DNA polymerase p 530-550 -DNA polymerase is stationary -primed by RNA fragments -extended in 5’--> 3’ direction -lagging strand loops out (coated with SSB) -lagging strand synthesis is discontinuous -leading strand synthesis is continuous http://www.youtube.com/watch?v=4jtmOZaIvS0
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DNA replication - DNA polymerase p 530-550 Chemistry: -nucleophilic attack of 3’-OH on α phosphate of dNTP -highly processive Structure: -highly conserved
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DNA replication - proofreading p 530-550 part of most DNA polymerases: an exonuclease located >25Å from polymerase site -error rate without proofreading = ~1/1,000 -error rate with proofreading = ~1/1,000,000 -error rate with DNA repair = ~1/100,000,000
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DNA replication - fnishing p 530-550 RNaseH
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This note was uploaded on 02/25/2010 for the course CHEM 4611 at Colorado.

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LECTURE 10 - DNA replication - overview Semiconservative...

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