DNA Replication

• Requirements of DNA

  Accurate

  • 1 copying error per million

  Fast

  • 1000 nt/sec in E coli.

  Capable of fixing errors

  Capable of bypassing DNA distortions
• Replication in Prokaryotes

  Single Replicon (Unit of replication)

  The two methods are theta replication and rolling circle replication
• Theta Replication

  Circular bacterial chromosomes and most plasmids

  dsDNA unwinds at the origin

  Replication bubble

  Replication forks
• Rolling Circle Replication


  Nick one nucleotide strand

  Add new nucleotides to the 3' end of the broken strand

  May go around the circle several times

  Linear DNA cleaved

  -Circularizes

  -Serves as template for complementary strand
• Requirements of Replication

  A single-stranded DNA template

  dNTPs

  Enzymes and Proteins
• Direction of Replication

  DNA polymerase can only add nucleotides to the 3' end of a strand

  DNA is always synthesized 5' to 3'
• Leading Strand

  Based on the 3' to 5' template strand

  Synthesized continuously
• Lagging Strand

  Based on the 5' to 3' template strand synthesized discontinuously w/ Okazaki fragments
• Mechanism of Bacterial DNA replication

  Initiation, Elongation, and Termination
• Initiation

  Single Replication Origin

  E. coli- OriC- 245 critical bps

  Initiator protein binds to the origin

  -Short Stretch of DNA unwinds
• Helicase

  Bind to lagging strand ssDNA

  Move 5' to 3' on this strand

  -Move replication fork

  Break Hydrogen bonds between base pairs of dsDNA

  Single Stranded Binding Proteins

  -Prevent ssDNA fromre-annealing
• Primase

  Synthesize RNA primers

  Complexed with DNA polymerase on the lagging strand
• Leading Strand

  One primer at 5' end
• Lagging Strand


  One primer for each Okazaki fragment