DNA Replication and Repair

The Central Dogma of Molecular Biology

DNA Replication

• genetic information is passed on to the next generation

• semi-conservative

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Parent molecule with two complementary

moleculesParental strands

separateEach parental strand

is a template

Each daughter DNA molecule consists of one

parental and one new strand

Overview of replicationInitiation

• DNA is unwound and stabilized• Origins of replication: Replication bubble and replication fork

Priming

• RNA primers bind to sections of the DNA and initiate synthesis

Elongation• Leading strand (5’ 3’) synthesized continuously• Lagging strand synthesized discontinuously then fragments are joined• RNA primer replaced by DNA

Proofreading

• Mismatch repair by DNA polymerase• Excision repair by nucleases

Review of DNA structure

• double helix• each strand has a 5’

phosphate end and a 3’ hydroxyl end

• strands run antiparallel to each other

• A-T pairs (2 H-bonds), G-C pairs (3 H-bonds)

STEP 1 Initiation at origins of replicationseparation sites on DNA strands

• Depend on a specific AT-rich DNA sequence– Prokaryotes – one site– Eukaryotes – multiple sites

• Replication bubble• Replication fork• Proceeds in two directions from point of origin

The proteins of initiation1. Helicase –

unwinds double helix

2. Single-strand binding proteins – holds DNA apart

STEP 2 Priminginitiation of DNA synthesis by RNA

RNA primers bind to unwound sections through the action of primase– leading strand –

only 1 primer– lagging strand –

multiple primers– replaced by DNA

later

STEP 3Elongation of a new DNA strandlengthening in the 5’ 3’ direction

DNA polymerase III can only add nucleotides to the 3’ hydroxyl end

Leading strand- DNA pol III – adds nucleotides

towards the replication fork; - DNA pol I - replaces RNA with DNALagging strand- DNA pol III - adds Okazaki

fragments to free 3’ end away from replication fork

- DNA pol I - replaces RNA with DNA- DNA ligase – joins Okazaki

fragments to create a continuous strand

STEP 4Proofreadingcorrecting errors in replication

Mismatch repair• DNA pol III – proofreads

nucleotides against the template strand

Excision repair• nuclease – cuts damaged segment• DNA pol III and ligase – fill the gap

left

Telomeres at 5’ ends of lagging strands

• no genes, only 100 – 1000 TTAGGG sequences to protect genes

• telomerase catalyzes lengthening of telomeres