DNA replication Flashcards
semi-conservative
after replication each double helix contains one parent strand and one new daughter strand
bidirectional
replication simulataneously moves away from origin in both directions
Okazaki fragments
short fragments formed on the lagging strand
origin of replication
one site per chromosome in prokaryotes, and 100’s in eukaryotes
fork
sites at which DNA synthesis is occuring
origin binding protiens
recognize and bind to origins or replication (AT rich sequences)
helicases
unwinds the helix ahead of the replication fork
ssbp
after the helicase unwinds the strands, ssbp bind to each single strand to keep them separated
primase
catalyzes the rxn forming the RNA to serve as a primer for the polymerases
DNA polymerase I
in prokaryotes
less processivity than III. has a clean-up fxn. mediates replacement of primer with 5-3 exonuclease and 5-3 polymerase activity. proofreads 3-5
DNA polymerase III
in prokaryotes
the major replicative enzyme. Higher processivity b/c it has a sliding clamp which keeps it attached to DNA over long distances
DNA Ligase
binds the Okazaki Fragments/segments of lagging strand.
Topoisomerase/gyrase
act to prevent extreme supercoiling of parental helix, by breaking and rejoining DNA. Gyrase- mostly foudn in prokaryotes and inhibited by quinolones.
Telomerase/reverse transcriptase
Retroviruses- synthesize DNA from RNA.
Telomerase- has reverse transcriptase activity. it carries it’s own RNA template to restore telomeres in human cancer cells and stem cells. Telomoerase is supressed in normal cells.
how polymerase creates phosphodiester bonds during addition of dNTP
adds dexyribonucleotides to the 3’ hydroxyls of the RNA primer, and then to the growing strand.
DNA polymerase requires ? to start and only synthesizes in a ? direction, and it corrects replication errors by ? activity.
RNA primer, 5-3, 3-5 exonuclease proofreading
Order of events
origin binding protiens bind, helicase unwinds, ssbp and topoisomerases maintain openness, primer binds, polymerase synthesized. okazaki fragments binded by ligase. RNA primers removed and replaced by DNA polymerase I.
Leading strand
continuous copying in 3-5 direction towards replication fork
lagging strand
discontinuous copying in 3-5 direction away from replication fork.
End Replication Problem
Lagging stand cannot be synthesized to very end, b/c you need an RNA primer to bind upstream from each synthesized region. There is nowhere to bind to synthesize the end. So the end gets shorter and shorter until it signals cell death.
Eukaryotes have 3 polymerases
Pol-a, a holoenzyme has primase and polymerase activity. It starts synthesis for about 20dNTP and then is swapped for Pol-g on lagging strand, and Pol-e on leading strand.
Fidelity of replication is high b/c
polymerases discriminate based on H-bonds between base pairs, and geometry of bonds. Errors during replication are corrected by 3-5 proofreading.
