Semiconservative Replication
DNA Replication
Chapter 11
Wikipedia
Kornberg’s DNA Polymerase I DNA Polymerase Reaction
Requirements for DNA synthesis DNA Polymerase Mg++ dNTP single stranded DNA template Priming 3’ OH Consider how the active site descriminates among the dNTP’s.
Other Enzymatic Activities Origin of Replication
E. coli DNA Polymerases 9 mers- TTATTTCCAC Enzymatic Activities I II III . 5’-3’ synthesis + + + Pair 13mers GATCTCTTATTAG
Required Primer + + + Order of Interaction 1. dnaA binds 9mers 3’-5’ Exonuclease + + + 2. dnaBC bind dnaA and 13mer -dnaB ATP dependent (Proofreading Activity) helicase 3. ssbp bind unwound DNA stabilizing it 5’-3’ Exonuclease 4. Gyrase (topoisomerase) relieves torsional stress (Replacement Activity) + - - . 5. Primase (dnaG) synthesizes short primers 6. DNA polymerase III initiates synthesis
DNA Polymerase III mutation lethal – essential to replication
DNA Polymerase I mutation viable – higher rate of mutation
DNA Polymerase II mutation viable – function unknown
1 dnaA Discontinuous Replication
DNA Polymerase synthesizes 5’-3’ only
http://oregonstate.edu/instruction/bb492/figletters/FigH3.html
Two Replication Forks Replication Fork
http://oregonstate.edu/instruction/bb492/figletters/FigH2.html
DNA Polymerase Clamp Replication Fork
Improved Processivity http://www.wehi.edu.au/education/wehi-tv/dna/replication.html
2 Simultaneous Synthesis Eukaryotic Issues
• Size of Genome • Ends of Linear Chromosomes
Movie
Size of Eukaryotic Genome Multiple Origins of Replication
Genome Size – E coli 4.6 Mbp genome – Human 3330 Mbp genome (Haploid)
Speed of Polymerase E. coli 100kb/min with two replication forks 23 min to copy genome
Humans 1kb/min with 2 replication forks per chromosome http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/D/DNAReplication.html 47.7 days to replicate genome
typical S phase – 2-3 hours embryonic S phase 3 minutes Eukaryotes can vary origins during development
Problem with linear chromosomes Telomerase
http://www.steve.gb.com/images/science/telomerase.png
3 Telomerase Biology
• Aging and Senescence
• Telomerase and Cancer
http://learn.genetics.utah.edu/features/telomeres/
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