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ibs602_rodgers_1_opt

ibs602_rodgers_1_opt - IBS 602 David Rodgers BBSRB B269...

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DNA replication IBS 602 David Rodgers BBSRB B269 257-5205 [email protected] outline replication origin and initiation primer formation replication/elongation termination cell division, complications first prokaryotic (E. coli) then eukaryotic sources: Molecular Biology of the Gene, Watson et al. (Ch. 7-8) Biochemistry, Voet and Voet (Ch. 31) Molecular Biology of the Cell, Alberts et al. (Ch. 4, Ch. 5) Genes VII, Lewin (Ch. 12,13) DNA Replication, Kornberg and Baker
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DNA replication reviews: Donachie and Blakely (2003) Coupling the initiation of chromosome replication to cell size in Escherichia coli. Curr. Opin. Microbiol. 6, 146. Mott and Berger (2007) DNA replication initiation: mechanisms and regulation in bacteria. Nat. Rev. Micro. 5, 343. Sherratt (2003) Bacterial chromosome dynamics. Science 301, 780. Pelizon (2003) Down to the origin: Cdc6 protein and the competence to replicate. Trends Cell Biol. 13, 110. Mendez and Stillman (2003) Perpetuating the double helix: molecular machines at eukaryotic DNA replication origins. BioEssays 25, 1158. Gilbert (2004) In search of the holy replicator. Nat. Rev. 5, 1.
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DNA replication basic reaction chapter 2 of the Watson text describes some of the classic experiments in the field
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origin/initiation E. coli chromosome -- 4.6 million base pairs oriC -- replication origin 9 bp repeat consensus TTA/TTNCACA DnaA protein -- recognizes 9 bp repeat in oriC -- binds ATP tightly and hydrolyzes slowly in a DNA dependent manner -- oligomerizes in ATP bound form Fujikawa et al. (2003) Nuc. Acid Res. 31, 2077-2086. Erzberger et al. (2006) Nat. Struct. Mol. Biol.
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origin/initiation oriC -- 14 GATC sequences -- methylation sites
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origin/initiation DnaA complex melts 13bp repeat sites 5'-GATCTnTTnTTTT-3' -- requires bound ATP -- must be above a minimum temperature -- approx. 45bp open single strand binding proteins (SSB) and DnaB bind and unwind more duplex --DnaC released negative/positive supercoiling
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origin/initiation DnaB helicase -- separates strands -- ATP hydrolysis -- 5'-->3' -- acts as a hexamer Singleton et al. (2000) Cell 101, 589.
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