MBIO


DNA polymerase


looks like a right handpalm made of beta sheets and is primary catalytic sitefingers bind incoming nucleotides and trap the correct dNTPthumb stabilizes reaction and processivity by maintaining interaction with the most recent dNTP


what are the 2 ions used by polymerase to stabilize


MG++ and ZN++ first removes a hydrogen and creates a nucleophilic attack on alphasecond maintains the beta and gamma phosphates


Operation of exonucleases


error occurssynthesis stops or slowsexonuclease removes mismatchresume synthesisonly 1 error in 10 to the 10 bps


addidtion of RNA primers


RNase makes nick in primerexonuclease removes primerprimer template junction exposedpolymerase fills junctionnick joined by ligase(primers must be removed from nascent DNA)


function of Topoisomerase 2


polymerase reaches super coiltopoisomerase2 binds and creates either 1 or 2 nicks in DNADNA uncoils and reanneals allowing synthesis to continue


polymerase alpha:


same as primase in prokaryotes:adds RNA primers


Polymerase S or E


they slide with the DNA clamp to extend DNA clamping down the DNA pol added at start site of primer and synthesizes DNA untill reaching next primer and falling off


Start of Replication


addition of ATP to the clamp loader to activate it by opening itbind the sliding clamp to the loaderbegins synthesis of DNAhydrolize the ATP on the loader to dessociate the loader


DNA replication at the fork


consists of the polymerase core, the T protein with flex linkers, the gamma complex (clamp loader), and sliding clampleadin strand is proccessive and is synthesized continuously from one side to the otherlagging strand rna primers are added, okazaki fragments are completed, dna pol is released, clamp is loaded on to newly primed lagging strand to synthesize more lagging strands, helicase accosiates with polymerase


ecoli initaition of replication (prokaryote)


dnaA and atp mind to originmany dnaAs denature dnahelicase (dnaB) bound by helicase loader (dnaC)helicase denatures dnaprimase bound and adds primerspolymerase added (holoenzyme) slidding clamp loadeddna synthesized


eukaryotic replication


ORC binds to replication originCdc6 and Cdt1 bindMcm2-7 complex addedpol B and E replace Cdc6 and Cdt1pol A added thus creating primasesliding clamp and clamp loader added (PCNA+RF-C)initiation of lagging strand synthesis


CDK activity


high- existing pre-RC activation/new formation inhibittedlow- pre-RC formation allowed/ no preRC activation


mitosis phases


G1- CDK levels LowS- CDK levels high


resolving dna end


3' hang telomerase adds primer to 3'synthesizes 5'repeates many times to extend dnasynthesizes rest of 3'ends once inhibitor proteins cover dna ensmall over hang still persists