DNA Replication Mechanism Quiz

DNA Replication

• The process of copying the entire genome prior to cell division
• Ensures the integrity of the genome across generations
• Occurs during the S phase of the cell cycle in eukaryotes
• Slower replication rate in eukaryotes results in higher fidelity/accuracy
• Humans add 3000 nucleotides per minute; bacteria add 30,000 per minute
• Relies on the complementarity of DNA strands, following the AT/GC rule

Mechanism of DNA Replication

• Initiation:

  • Initiator proteins bind to specific base sequences on DNA called "sites of origin"
  • Prokaryotes have a single origin site (e.g., oriC in E. coli)
  • Eukaryotes have multiple sites of origin (e.g., ARS sequences in yeast)
  • Helicase unwinds and separates the double-stranded DNA, creating a replication fork
  • Single-Strand Binding Proteins stabilize the separated strands
  • Topoisomerase relieves stress on the DNA molecule during separation

• Elongation:

  • Synthesis always occurs in the 5´-3´ direction
  • RNA primers are required to initiate nucleotide addition
  • Primase synthesizes the RNA primer
  • DNA polymerase adds nucleotides to the 3´ end of the primer
  • Leading strand is synthesized continuously
  • Lagging strand is synthesized discontinuously in short segments called Okazaki fragments
  • Okazaki fragments are ~1000-2000 bp in prokaryotes and ~100-200 bp in eukaryotes
  • DNA ligase joins the Okazaki fragments together

• Termination:

  • In prokaryotes, replication forks reach specific "termination sites"
  • In eukaryotes, telomerase solves the problem of gaps left after RNA primer removal at the 5´ end of the lagging strand

Proteins Involved in DNA Replication

• DNA polymerase: Adds nucleotides to the growing DNA strand. Proofreads for errors.
• Helicase: Unwinds the DNA double helix
• Topoisomerase: Relieves torsional stress from unwinding
• RNA primase: Synthesizes RNA primers
• Single-strand binding proteins: Prevents reannealing of separated DNA strands
• DNA ligase: Joins Okazaki fragments on the lagging strand

Characteristics of DNA Polymerase

• Synthesizes DNA in only one direction (5´-3´)
• Requires a pre-existing double-stranded DNA to begin synthesis
• Cannot initiate DNA synthesis from scratch

Proofreading and Error Correction

• DNA polymerase initially makes errors at a rate of 1 in 10,000 base pairings
• Proofreading enzymes correct these errors
• DNA polymerases possess 3´-5´ exonuclease activity, allowing them to remove mismatched nucleotides
• This process improves accuracy to 1 in 1 billion base pairing errors

Replication Bubbles

• Replication occurs in both directions along the DNA
• Both strands are replicated simultaneously
• This process generates replication bubbles

Telomeres

• Telomerase extends the lagging strand's 5´ end, preventing degradation
• This solves the problem of gaps left after RNA primer removal
• Essential in eukaryotes for maintaining chromosome stability