Chapter 6 - Replication Repair Recombination cell lives...

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1 Replication, Repair & Recombination cell lives in “chaotic environment” lots of chemical stress leads to modification of DNA = mutations need to ensure that the integrity/fidelity of the genome remains high & that cell can donate an exact replica to daughter cells = Repair, Replication and Recombination • double stranded DNA template ideally suited to ensure fidelity of transmission - base-pairing & template dictate mechanisms. DNA’s 3 R’s Base-pairing is a key feature! germ cell versus somatic cells … Replication - semi-conservative - each strand serves as a template for the other one - base pairing rules dictate which nt to insert - ensures speed (~ 8 hrs) and accuracy (10 -7 to 10 -9 errors)
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2 Origin(s) of Replication cell uses replication machinery (protein complex) to synthesize new strands very efficient, very accurate Process includes: 1. strand separation - lots of weak H-bonds 2. ID origins - small stretches DNA (AT rich) that can be opened easily [ one in bacteria; 10,000 in humans] 3. Initiator Proteins - open helix, increase access for replication machinery 4. Bidirectional movement to synthesize new DNA Replication Machinery - keeps strands separate for short stretches; also close together rate of extension approx. same both strands requires energy - hydrolysis of NTP to NMP plus release of PPi (into DNA strand) e.g. CTP PPi 2x[Pi] CMP
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3 06_10_5prime_3prime.jpg 2x[Pi] 1. base pairing dictates which nt as H-bonds form. 2. the 3’ -OH group of ribose attacks 5’ end of incoming nt, 3. releases PPi as sugar- phosphate bond forms 4. hydrolysis of PPi drives reaction forward (irreversibly) 1 DNA polymerase: • key enzyme of replication machine • adds nt one at a time to 3’ end of a growing strand synthesizes DNA is 5’ to 3’ direction ONLY! • high processivity - stays on DNA strand for multiple rounds of nt addition asymmetric replication fork: this strand (LEADING) can be continuously synthesized this strand (LAGGING) must have discontinuous synthesis Lagging strand - a ‘backstitching,’ discontinuous synthesis of new strand - creation of a series of Okazaki Fragments
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4 06_12_asymmetrical.jpg Okazaki Fragments started here DNA polymerase - proofreading • fairly accurate but some mismatch of base pairing is possible
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