LECTURE 4&5 - The DNA replication fork collaborate or die-Other replication factors it takes a v

LECTURE 4&5 - The DNA replication fork collaborate or die-Other replication factors it takes a v

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Unformatted text preview: DNA polymerase summary 1. DNA r eplication is semi-conser vative. 2. DNA polymer ase enzymes ar e specialized for differ ent functions. 3. DNA pol I has 3 activities: polymer ase, 3-->5 exonuclease & 5-->3 exonuclease. 4. DNA polymer ase str uctur es ar e conser ved. 1. But: Pol cant star t and only synthesizes DNA 5-->3! 1. Editing (pr oofr eading) by 3-->5 exo r educes er r or s. 2. H igh fidelity is due to the r ace between addition and editing. 3. M ismatches disfavor addition by DNA pol I at 5 successive positions. The er r or r ate is ~ 1/10 9 . Replication fork summary 1. DNA polymer ase cant r eplicate a genome. Pr oblem Solution ATP? No single str anded template H elicase + The ss template is unstable SSB (RPA (euks))- No pr imer Pr imase (+) No 3-->5 polymer ase Replication for k Too slow and distr ibutive SSB and sliding clamp- 2. Replication for k is or ganized ar ound an asymmetr ic, DNA- polymer ase I I I dimer . 3. Both str ands made 5-->3. 4. Leading str and is continuous; lagging str and is discontinuous. DNA polymerase cant replicate a genome! 1. No single str anded template 2. The ss template is unstable 3. No pr imer 4. No 3-->5 polymer ase 5. Too slow and distr ibutive Solution: the replication fork 1. No single-str anded template 2. The ss template is unstable 3. No pr imer 4. No 3-->5 polymer ase 5. Too slow and distr ibutive Schematic drawing of a replication fork DNA polymerase holoenzyme QuickTime and a DV - PAL decompressor are needed to see this picture. DNA replication factors were discovered using temperature sensitive mutations 1. No single str anded template 1. The ss template is unstable 2. No pr imer 3. No 3-->5 polymer ase. 4. Too slow in vitr o. M utations that inactivate the DNA r eplication machiner y ar e lethal. Temper atur e sensitive (conditional) mutations allow isolation of mutations in essential 37 C 42 C 42 C, M utant gene over expr essed A hexameric replicative helicase unwinds DNA ahead of the replication fork 1. No single str anded template 1. The ss template is unstable 2. No pr imer 3. No 3-->5 polymer ase....
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LECTURE 4&5 - The DNA replication fork collaborate or die-Other replication factors it takes a v

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