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DNA Replication AP Biology 2007-2008 Watson and Crick AP Biology 1953 article in Nature Double helix structure of DNA “It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic AP Biology material.” Watson & Crick Directionality of DNA You need to PO4 nucleotide number the carbons! it matters! N base 5 CH2 This will be IMPORTANT!! O 4 3 AP Biology 1 ribose OH 2 The DNA backbone Putting the DNA backbone together refer to the 3 and 5 ends of the DNA the last trailing carbon Sounds trivial, but… this will be IMPORTANT!! 5 PO4 base 5 CH2 O 4 1 C 3 O –O P O O 5 CH2 2 base O 4 1 2 3 OH AP Biology 3 Anti-parallel strands Nucleotides in DNA backbone are bonded from phosphate to sugar between 3 & 5 carbons 5 3 3 5 DNA molecule has “direction” complementary strand runs in opposite direction AP Biology Bonding in DNA 5 hydrogen bonds 3 covalent phosphodiester bonds 3 5 ….strong or weak bonds? AP Biology How do the bonds fit the mechanism for copying DNA? Base pairing in DNA Purines adenine (A) guanine (G) Pyrimidines thymine (T) cytosine (C) Pairing A:T 2 bonds C:G 3 bonds AP Biology Copying DNA Replication of DNA base pairing allows each strand to serve as a template for a new strand new strand is 1/2 parent template & 1/2 new DNA AP Biology DNA Replication Let’s meet the team… Large team of enzymes coordinates replication AP Biology Replication: 1st step Unwind DNA helicase enzyme unwinds part of DNA helix stabilized by single-stranded binding proteins helicase single-stranded binding proteins AP Biology replication fork Replication: 2nd step Build daughter DNA strand add new complementary bases DNA polymerase III DNA Polymerase III AP Biology 5 Replication Adding bases can only add nucleotides to 3 end of a growing DNA strand need a “starter” nucleotide to bond to strand only grows 53 AP Biology B.Y.O. ENERGY! The energy rules the process 3 energy DNA Polymerase III energy DNA Polymerase III energy DNA Polymerase III DNA Polymerase III energy 3 5 Okazaki Leading & Lagging strands Limits of DNA polymerase III can only build onto 3 end of an existing DNA strand 5 3 5 3 5 3 5 5 5 Lagging strand ligase growing 3 replication fork Leading strand 3 Lagging strand Okazaki fragments joined by ligase AP Biology “spot 3 welder” enzyme 5 3 DNA polymerase III Leading strand continuous synthesis Replication fork / Replication bubble 3 5 5 3 DNA polymerase III leading strand 5 3 3 5 3 5 5 5 3 lagging strand 3 5 3 5 lagging strand 5 5 leading strand 3 growing replication fork leading strand 3 lagging strand 5 5 AP Biology growing replication fork 5 5 5 3 Starting DNA synthesis: RNA primers Limits of DNA polymerase III can only build onto 3 end of an existing DNA strand 5 3 3 5 5 3 5 3 5 growing 3 replication fork DNA polymerase III primase RNA 5 RNA primer built by primase serves as starter sequence DNA polymerase III AP for Biology 3 Replacing RNA primers with DNA DNA polymerase I removes sections of RNA primer and replaces with DNA nucleotides DNA polymerase I 3 5 5 5 3 ligase growing 3 replication fork RNA 5 3 But DNA polymerase I still can only build onto 3 end of an existing DNA strand AP Biology Replication fork DNA polymerase III lagging strand DNA polymerase I 5’ 3’ ligase primase Okazaki fragments 5’ 3’ 5’ SSB 3’ helicase DNA polymerase III 5’ 3’ leading strand direction of replication AP Biology SSB = single-stranded binding proteins DNA polymerases DNA polymerase III 1000 bases/second! main DNA builder Roger Kornberg 2006 DNA polymerase I 20 bases/second editing, repair & primer removal DNA polymerase III enzyme AP Biology Arthur Kornberg 1959 Editing & proofreading DNA 1000 bases/second = lots of typos! DNA polymerase I proofreads & corrects typos repairs mismatched bases removes abnormal bases repairs damage throughout life AP Biology reduces error rate from 1 in 10,000 to 1 in 100 million bases Fast & accurate! It takes E. coli <1 hour to copy 5 million base pairs in its single chromosome divide to form 2 identical daughter cells Human cell copies its 6 billion bases & divide into daughter cells in only few hours remarkably accurate only ~1 error per 100 million bases ~30 errors per cell cycle AP Biology Any Questions?? AP Biology 2007-2008