Theme 4 Module 2

• DNA is the macromolecule that determines the characteristics of the cell
• Watson and Crick proposed the characteristic helical structure of DNA and also proposed that the base-pairing in DNA should allow for a mechanism by which the genetic information in DNA could be copied
• Semiconservative model of DNA replication

  When a DNA double helix replicates, each of the two daughter DNA molecules would have one old strand, from the parental molecule and one newly made strand
• Meselson and Stahl's experiment to demonstrate semiconservative DNA replication

  1. Cultured E. coli in medium with 15N, then transferred to 14N medium

  2. Extracted DNA samples and centrifuged to separate based on density

  3. Observed hybrid band after one round of replication, then two distinct bands after many rounds
• Meselson and Stahl's results supported the semiconservative model of DNA replication
• DNA replication is similar in prokaryotes and eukaryotes, following the semiconservative model
• DNA replication process


  1. Begins at origins of replication

  2. Template strand copied from 3' to 5' end

  3. Daughter strand elongates 5' to 3'

  4. Complementary nucleotides bond to template and form phosphodiester bonds

  5. Replication forks form at origins
• Leading strand


  Continuous replication from a single primer, in the 5' to 3' direction
• Lagging strand


  Discontinuous replication, with Okazaki fragments formed from separate primers
• DNA polymerase and other proteins play important roles in the DNA replication process
• DNA replication


  Only adds nucleotides to the 3' end of a polymerizing DNA molecule
• DNA replication


  1. Replication from this strand requires that DNA polymerase replicate the DNA in a direction that is away from the replication fork

  2. Lagging strand contains segments or fragments of DNA called Okazaki fragments

  3. Only one primer is required for the synthesis of the leading daughter strand, each Okazaki fragment on the lagging strand is formed by separate primers

  4. DNA polymerase forms an Okazaki fragment, another DNA polymerase replaces the RNA primer sequences with DNA nucleotides

  5. Many other proteins serve important roles during the DNA replication process
• Replication complex


  • Various proteins that participate in DNA replication form a single large complex
  • DNA helicase enzymes bind to the parental DNA strands at the origin of replication and initiate the unwinding of the DNA double helix
  • Single-stranded binding proteins bind to and stabilize each parental strand until elongation can begin
  • Topoisomerases bind upstream of the replication fork and minimize the torsional strain from the unwinding at the replication fork
  • These proteins serve as initiator proteins that trigger the unwinding at the origin of replication
• Primer synthesis


  1. RNA primase synthesizes short RNA stretches of nucleotides complementary to the parental strands

  2. DNA polymerase III does most of the elongation work in prokaryotes, while DNA polymerase I removes the RNA primer and replaces it with DNA nucleotides
• Replication process is similar between prokaryotes and eukaryotes, but eukaryotes utilize a different set of DNA polymerases
• Lagging strand synthesis


  1. Synthesis of the lagging strand is delayed relative to the leading strand

  2. Each new fragment of the lagging strand cannot be replicated until enough of the template DNA is revealed at the replication fork

  3. In eukaryotes, the replacement of the RNA primer with DNA nucleotides leaves a sugar phosphate backbone at the 3' end with a free phosphate backbone

  4. DNA ligase joins the 3' end of a fragment to an adjacent DNA nucleotide by catalyzing the phosphodiester bond formation
• DNA replication accuracy


  • DNA polymerases proofread each added nucleotide relative to the template strand
  • If an incorrect nucleotide pairing is detected, DNA polymerase removes the incorrect nucleotide and adds the correct one
  • Mismatched nucleotides can sometimes evade the proofreading mechanism, and other enzymes can help correct replication errors
• Replication in eukaryotes originates at many origins of replication along the linear chromosomes, while in prokaryotes it begins at one origin of replication
• Eukaryotic DNA replication has a leading strand that replicates the whole template strand, but the lagging strand cannot be fully replicated due to the linear nature of eukaryotic chromosomes
• Telomeres


  Special nucleotide sequences at the ends of linear eukaryotic chromosomes, mainly made up of repetitions of one short nucleotide sequence
• Telomere length


  • Telomeres become shorter during successive rounds of replication in somatic cells
  • Telomere shortening does not occur in gametes or stem cells due to the presence of the telomerase enzyme
  • Telomerase is a reverse transcriptase that can elongate the telomere regions by adding telomere repeats