DNA Replication Overview

DNA Replication Overview

• DNA replication is the process of creating two identical copies of a DNA molecule from a single original DNA molecule.
• This process ensures genetic information is passed on correctly during cell division.
• The semi-conservative model, proven by the Meselson-Stahl experiment, shows each new DNA molecule consists of one original strand and one new strand.

Replication Models

• Semiconservative model: Each new DNA molecule contains one original strand and one newly synthesized strand.
• Conservative model: One DNA molecule retains both original strands, and the other consists of two newly synthesized strands.
• Dispersive model: Original and new DNA segments are interspersed in both new DNA molecules.

Meselson-Stahl Experiment

• This experiment, using isotopes of nitrogen (15N and 14N), confirmed the semiconservative model.
• E. coli were grown in a medium containing 15N, then transferred to a 14N medium.
• DNA extracted at various time points was analyzed using density gradient centrifugation.
• The results showed DNA progressively shifted to a lower density, supporting the semiconservative model.

Steps of DNA Replication

• Step 1: Strand Separation
  • Helicase unwinds the DNA double helix.
  • Topoisomerases relieve the tension created by unwinding.
  • Single-strand binding proteins prevent the separated strands from reannealing.
• Step 2: Building Complementary Strands
  • RNA primase synthesizes RNA primers, providing a starting point for DNA polymerase.
  • DNA polymerase III builds new DNA strands using complementary base pairing.
  • The leading strand is synthesized continuously, while the lagging strand is synthesized in short fragments (Okazaki fragments).
  • DNA polymerase I removes RNA primers and replaces them with DNA.
  • DNA ligase joins Okazaki fragments.
• Step 3: Dealing with Errors
  • DNA polymerase proofreads the newly synthesized DNA.
  • Mismatch repair systems identify and correct errors.
    • Recognition, excision of the error, gap-filling and ligation are some of the steps

DNA Replication in Prokaryotes and Eukaryotes

• Prokaryotes (bacteria) have a single replication origin, forming a replication bubble that expands bidirectionally.
• Eukaryotes have multiple replication origins, resulting in numerous replication bubbles that eventually merge.
• Eukaryotes use telomeres (repetitive sequences at chromosome ends) to prevent loss of genetic material during replication. Telomerase re-extends the telomeres.
• Eukaryotes have more variations of DNA polymerases compared to prokaryotes.

Key Enzymes

• Helicase: Unwinds the DNA double helix.
• Topoisomerases: Relieve tension caused by unwinding.
• Single-strand binding proteins (SSBs): Prevent reannealing of separated strands.
• RNA primase: Synthesizes RNA primers.
• DNA polymerase III: Builds new DNA strands.
• DNA polymerase I: Removes RNA primers and replaces them with DNA.
• DNA ligase: Joins Okazaki fragments.
• DNA polymerase II: Repairs DNA damage.

Rate of Replication

• In eukaryotes, new DNA is synthesized at a rate of approximately 50 base pairs per second at each replication fork.