mbb 331 lecture 8 (thurs feb 6)

DNA Damage and Repair

Overview

Topics covered:

  • DNA mutations

  • DNA damage

  • DNA damage repair

Key Concepts

  • DNA Mutation: Permanent change in the DNA sequence, which can alter gene function and lead to various genetic disorders.

  • DNA Damage: Physical or chemical alteration of DNA that can result from internal and external factors, impacting genetic integrity.

  • DNA Damage Repair Mechanisms: Complex processes involving various pathways that correct mutations and damages to maintain genetic stability, crucial for preventing diseases such as cancer.

Types of DNA Damage

  • Categories of DNA Damage:

    • Base alterations: Changes to individual bases that can lead to incorrect base pairing during replication.

    • Structural distortions: Changes resulting in altered DNA structure, such as bending, kinking, or other conformational changes that can impede replication and transcription.

Causes of DNA Damage:

  • Deamination: The loss of an amine group from a nucleotide, leading to mispairing during DNA replication.

  • Oxidative damage: Damage caused by reactive oxygen species (ROS) that can result in base modifications and strand breaks.

  • Damage by alkylation: Addition of alkyl groups to nucleotides, primarily affecting guanine, leading to mispairing and mutations.

  • UV irradiation: Exposure to ultraviolet light induces the formation of pyrimidine dimers, disrupting normal base pairing.

  • Errant replication and recombination: Mistakes during DNA replication or recombination events can introduce errors into the genetic code.

Repair Mechanisms

  • Direct Repair: A unique method that repairs DNA without removing the altered base, allowing for efficient correction of specific lesions.

  • Base Excision Repair (BER): Targets small, non-helix distorting base lesions. The process includes recognition of the damaged base, excision, and replacement.

  • Nucleotide Excision Repair (NER): Repairs bulky DNA adducts and helix-distorting damage through incision, excision, and synthesis of a new DNA strand.

  • Mismatch Repair (MMR): Corrects mispaired bases shortly after DNA replication, using the methylation status to identify which strand to repair.

  • Recombination Repair: Essential for handling double-strand breaks, this mechanism uses homologous sequences to accurately repair the DNA.

DNA Fidelity and Replication Accuracy

  • Replication Accuracy: DNA polymerases exhibit high fidelity during replication, primarily due to proofreading functions, resulting in an error rate of ~7 × 10^-6 per division.

  • DNA Repair Mechanisms: Additional mechanisms further reduce these errors to approximately ~1 × 10^-10 per division, ensuring faithful genetic replication and stability.

Types of Mutations

  • Point Mutations: Refers to changes in a single nucleotide pair. Types include:

    • Transitions: Substitution of a purine for another purine or pyrimidine for another pyrimidine.

    • Transversions: Substitution of a purine for a pyrimidine or vice versa.

    • Potential outcomes of mutations:

      • Nonsynonymous (Missense): Results in a different amino acid that can alter protein function.

      • Nonsense: Creates a premature termination codon, leading to truncated proteins and loss of function.

      • Silent (Synonymous): No change in the amino acid sequence despite nucleotide alteration.

  • Small Indels: Refers to insertions or deletions of nucleotides, which can cause:

    • Frameshift mutations: Disruption of the reading frame, potentially altering all downstream amino acids.

    • In-frame mutations: Changes in a few amino acids without affecting the overall reading frame.

Triplet Expansion Diseases

  • Example: Fragile X Syndrome: Caused by the expansion of CGG repeats leading to more than 200 copies, distinctly resulting in cognitive impairment and other developmental issues.

DNA Damage Mechanisms

  • Specific Damage Types:

    • Deamination: Leads to erroneous base pairing by converting cytosine to uracil, thus causing potential mutations.

    • Depurination: The loss of purine bases (adenine or guanine) results in abasic sites that can lead to significant genetic instability.

    • Oxidative Damage: ROS can induce modifications to all four nucleotides, resulting in mutagenic effects.

    • Alkylation: The addition of alkyl groups to guanine can disrupt normal base pairing, increasing the likelihood of mutations.

    • Radiation Damage: UV light triggers the formation of pyrimidine dimers, while gamma or X-ray exposure can cause double-strand breaks, leading to serious genomic instability.

Repair Mechanisms in Detail

  • Direct Repair:

    • Alkyl transferases: Enzymes that fix alkylated bases by transferring the alkyl group away from the damaged site.

    • Photolyase: An enzyme that utilizes light energy to specifically repair UV-induced thymine dimers, restoring normal base pairing.

  • Base Excision Repair (BER):

    • Recognizes damaged bases through the action of DNA glycosylase, followed by the AP endonuclease creating a nick at the AP site.

    • Subsequently, DNA polymerase fills in the gap, and ligation of the strand occurs to seal the repair.

  • Nucleotide Excision Repair (NER):

    • Involves the recognition and incision of the damaged DNA strand by excinucleases, leading to the synthesis of new DNA using DNA polymerase, finally followed by ligation to restore DNA integrity.

  • Mismatch Repair (MMR):

    • Identifies mismatched bases using methylation status to determine which strand needs repair. This process involves key proteins like MutS and MutL to detect and correct replication errors.

Final Notes

Understanding these mechanisms is crucial for studying genetic diseases and developing therapeutic strategies aimed at correcting mutations and maintaining genomic integrity. Continued research in this field may pave the way for innovative treatments for various hereditary conditions.