Watson & Crick. DNA Replication. Directionality of DNA. Double helix structure of DNA. The DNA backbone. Anti-parallel strands
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1 Watson and Crick 1953 article in Nature Replication Double helix structure of Directionality of You need to number the carbons! it matters! P 4 nucleotide N base This will be IMPRTANT!! 4 ribose 1 It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material. Watson & Crick H 2 The backbone Putting the backbone together refer to the and ends of the the last trailing carbon Sounds trivial, but this will be IMPRTANT!! P 4 base 4 C P base Anti-parallel strands Nucleotides in backbone are bonded from phosphate to sugar between & carbons molecule has direction complementary strand runs in opposite direction H 2 1
2 Bonding in covalent phosphodiester bonds hydrogen bonds.strong or weak bonds? AP How Biology do the bonds fit the mechanism for copying? Base pairing in Purines adenine (A) guanine (G) Pyrimidines thymine (T) cytosine (C) Pairing A : T 2 bonds C : G 3 bonds Copying Replication of base pairing allows each strand to serve as a template for a new strand new strand is 1/2 parent template & 1/2 new semi-conservative copy process Replication Let s meet the team Large team of enzymes coordinates replication Replication: 1st step Unwind helicase enzyme unwinds part of helix I d love to be helicase & unzip your genes stabilized by single-stranded binding proteins helicase Replication: 2nd step Build daughter strand add new complementary bases polymerase III But Where s the We re missing ENERGY something! for the bonding! What? single-stranded binding proteins 2
3 Energy of Replication Where does for bonding usually come from? You remember ATP! Are there other ways nucleotides? to get You out of bet! it? We come with our own! Energy of Replication The nucleotides arrive as nucleosides bases with P P P P-P-P = for bonding bases arrive with their own source for bonding bonded by enzyme: polymerase III GTP TTP CTP ATP modified nucleotide And we leave behind a nucleotide! GMP TMP CMP ADP AMP ATP GTP TTP CTP Replication Adding bases can only add nucleotides to end of a strand need a starter nucleotide to bond to strand only grows B.Y.. ENERGY! The rules the process no to bond need primer bases to add on to kazaki Leading & Lagging strands Limits of polymerase III can only build onto end of an existing strand Lagging strand kazaki fragments joined by spot welder enzyme polymerase III Leading strand Lagging strand Leading strand continuous synthesis Replication fork / Replication bubble polymerase III 3
4 Starting synthesis: RNA primers Limits of polymerase III can only build onto end of an existing strand polymerase III primase Replacing RNA primers with polymerase I removes sections of RNA primer and replaces with nucleotides polymerase I RNA RNA RNA primer built by primase serves as starter sequence for polymerase III But polymerase I still can only build onto end of an existing strand Chromosome erosion All polymerases can only add to end of an existing strand polymerase I Houston, we have a problem! Telomeres Repeating, non-coding sequences at the end of chromosomes = protective cap limit to ~50 cell divisions polymerase III telomerase Loss of bases at ends in every replication chromosomes get shorter with each replication AP limit Biology to number of cell divisions? RNA Telomerase enzyme extends telomeres can add bases at end different level of activity in different cells high in stem cells & cancers -- Why? TTAAGGG TTAAGGG 5 Replication fork polymerase III polymerase I kazaki primase fragments 5 5 SSB direction of replication polymerase III 5 helicase SSB = single-stranded binding proteins polymerases polymerase III 1000 bases/second! main builder polymerase I 20 bases/second editing, repair & primer removal polymerase III enzyme Thomas Kornberg?? Arthur Kornberg
5 Editing & proofreading 1000 bases/second = lots of typos! polymerase I proofreads & corrects typos repairs mismatched bases removes abnormal bases repairs damage throughout life 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 What does it really look like? Any Questions??
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