DNA Replication

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DNA Replication

• The reason why most of us dont have three eyes.

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The Three Requirements of Genetic Material

• Must be able to replicate.
• Must control the expression of traits.
• Must be able to change in a controlled way.

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DNA Replication

• What is DNA Replication?
• How fast does DNA replicate?
• How does DNA replicate?
• Whats amazing about DNA replication?

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Conservative replication Semiconservative
replication Dispersive replication
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DNA Replication

• Three Models
• Conservative replication an entirely new double
  stranded DNA molecule is produced
• Semi-conservative replication each strand
  serves as a template for assembly of a new
  complementary strand
• Dispersive replication breakage in both
  parental strands and reassembly

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Meselson-Stahl Experiment

• 1957, Matthew Meselson, Franklin Stahl,
  California Inst. Of Technology
• Heavy nitrogen 15N
• Shift to 14N
• Determine buoyant density in CsCl gradient

Time
DNA replicates in a semiconservative fashion
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What is DNA Replication?
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How Fast is DNA Replication?

• Facts
• In prokaryotes (E. coli) about 1000 bases per
  second.
• E. colis genome is 4.7 X 106 bases
• In eukaryotes - about 50 nucleotides per second.
• Average human chromosome has 150 X 106 bases.
• Answer
• Pretty darn fast?
• Not fast enough?

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Proteins Involved in DNA Replication

• Presynthesis
• Topoisomerase
• Single-stranded binding proteins
• Helicase
• Synthesis
• Primase
• Polymerase
• Exonuclease
• Ligase

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http//www.johnkyrk.com/DNAreplication.html
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Presynthesis Replication Proteins

• Topoisomerase - an enzyme responsible for
  creating a nick in a strand of supercoiled DNA,
  relaxing the helix. There are four types that
  fall into two main groups.
• Helicase - an enzyme that unwinds DNA.
• Single-stranded binding proteins attaches to
  the unwound single strand of DNA to stabilize it.
  Speeds up DNA replication.

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Proposed 3-D Models of Topoisomerase I and II
from the Protein Data Bank.
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DNA Replication Requires Enzymes

• Primase an enzyme that attaches an RNA primer
  to the single stranded DNA.
• DNA Ligase forms a covalent link between the 3
  OH and 5 phosphate groups

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DNA Primase

• Role synthesis of short RNA primers on single
  stranded DNA templates (every 500 2,000
  nucleotides on the lagging strand).
• Three domains
• N-terminal zinc-binding motif.
• Central region with RNA polymerase sequence
  homology.
• C-terminal residue that interacts with helicase.

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DNA Polymerase
Synthesis Proofreading Error Correction
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DNA Polymerase

• DNA Polymerase I (Pol I) first enzyme discovered
  with polymerase activity, best characterized
  enzyme
• 5' to 3' elongation (polymerase activity)
• 3' to 5' exonuclease (proof-reading activity)
• 5' to 3' exonuclease (repair activity)
• DNA Polymerase II (Pol II)
• Repairs gaps, 5 3
• DNA Polymerase III
• Provides the primary 5 to 3 polymerase activity
• Slight exonuclease activity.

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DNA Replication Summary

• http//www.ncc.gmu.edu/dna/replicat.htm