DNA Repair '

Overview of DNA Damage and Repair Mechanisms

  • DNA experiences damage from multiple sources including:

    • X-rays
    • Alkylating agents
    • Oxygen radicals
    • UV light
    • Polycyclic aromatic hydrocarbons

  • DNA repair mechanisms must effectively address several types of damage including:

    • Replication errors
    • Uracil incorporation
    • Abasic sites
    • Bulky adducts
    • 6-4 Photoproducts (6-4PP)
    • 8-Oxoguanine
    • Single-strand breaks
    • Interstrand cross-links
    • Double-strand breaks

DNA Repair Processes

Types of DNA Repair Mechanisms

  • Direct Repair

  • Base Excision Repair (BER)

  • Nucleotide Excision Repair (NER)

  • Mismatch Repair (MMR)

  • Homologous Recombination Repair (HR)

  • Non-Homologous End Joining (NHEJ)

  • Questions to Consider:

    • Describe the process of repair by > Direct reversal, Base excision repair, Nucleotide excision repair, Homologous recombination repair, Non-homologous end joining.
    • Identify what type of damage is repaired in each mechanism.

Importance of DNA Repair

  • Despite the efficiency of DNA polymerase, some mutations persist due to incorrect repair mechanisms.
  • Cellular damage to DNA from biological and environmental factors necessitates robust DNA repair processes.
  • Review material from previous lectures on related topics, specifically Lectures 17 & 18.

Direct Repair Mechanisms

  • Direct Repair restores bases to their original state without removing them.
    • Thymine Dimer Repair:
    • DNA photolyase cleaves the bonds between thymine dimers, restoring the normal structure of thymines.
  • Methylated Base Repair:
    • Alkyltransferase removes the methyl group from 6-Methylguanine, restoring correct guanine structure.

Base Excision Repair (BER)

  • Mechanism in Bacteria (e.g., E. coli):

    1. DNA N-glycosylases cut out the damaged base.
    2. AP endonuclease recognizes the missing base and cuts the DNA backbone.
    3. DNA polymerase I removes the base-less nucleotide and replaces it with the correct one.
    4. DNA ligase seals the DNA backbone.

  • Mechanism in Eukaryotes:

    • DNA polymerase β can directly remove the base-less nucleotide and fill in the gap, followed by action from DNA ligase.
    • Alternatively, DNA polymerase δ or ε synthesizes a short segment, followed by removal of the flap by flap endonuclease and sealing by DNA ligase.

Nucleotide Excision Repair (NER)

  • Process: Removes several nucleotides from the damaged strand.
  • Clinical Relevance:
    • Xeroderma Pigmentosum is cancer-prone due to mutations in one of seven genes coding for NER proteins, leading to skin cancer due to DNA repair deficiency.

Mismatch Repair (MMR)

  • Function: Removes mispaired bases in DNA.
  • Error Rate: DNA polymerase mispairs at about 1 in 100 million nucleotides.
  • Mechanism in Bacteria:
    1. Mut proteins scan for mismatches in DNA.
    2. MutS/MutL complex binds to MutH at hemimethylated sites.
    3. MutH cuts the non-methylated strand.
    4. The non-methylated strand is digested until the mismatch is reached, then DNA polymerase fills in the gap and DNA ligase seals it.

Double-Strand Break Repair

Homologous Recombination (HR)

  • Occurrence: This process happens during late S-phase and G2 phase.
  • Steps:
    1. Exonuclease generates single-stranded DNA to allow for strand invasion.
    2. DNA polymerase fills in gaps, and endonuclease cuts the DNA backbone.
    3. Ligation occurs to seal any nicks.
  • Advantage: Highly accurate due to the use of sister chromatids as templates, preventing sequence loss during repair.

Non-Homologous End Joining (NHEJ)

  • Timing: Occurs during the G1 phase.
  • Steps:
    1. Proteins bind to and bridge broken ends of the DNA.
    2. Ends of the DNA are processed and rejoined.

Summary of Other Types of Nucleases

  • Exonuclease: Removes nucleotides from the ends of DNA fragments.
  • Endonuclease: Breaks an internal bond between the sugar and phosphate groups of the backbone.

Reflection Questions

  • What components of the course were most and least helpful for your learning?
  • What suggestions do you have for improving the course format for future students?