Chapter 15: DNA and the Gene: Synthesis and Repair

Chapter 15: DNA and the Gene: Synthesis and Repair

Overview of DNA Replication

• Electron Micrograph: Illustrates DNA replication with original DNA double helix diverging at the replication fork into two helices.
• Replication Fork: Area where DNA synthesis occurs, indicating active replication of the DNA strands.

15.1 Experimental Evidence for DNA as Hereditary Material

15.1.1 Hershey-Chase Experiment Summary

• Research Question: Do viral genes consist of DNA or protein?
  • Hypotheses:
  • DNA Hypothesis: T2 virus genes consist of DNA.
  • Protein Hypothesis: T2 virus genes consist of protein.

Experimental Setup

1. Labeled Viruses:
   • Set of T2 viruses grown in radioactive $^{32}P$ (present in DNA).
   • Another set grown in radioactive $^{35}S$ (present in protein).
2. Infection: Viruses infect separate cultures of E. coli.
3. Agitation: Cultures agitated in a blender to separate capsids from cells.
4. Centrifugation: Solutions separated to locate where radioactivity exists.

Predictions

• DNA Hypothesis: Radioactive DNA will be found in the bacterial pellet.
• Protein Hypothesis: Radioactive protein will be found in the pellet.

Results

• Radioactive DNA was in the pellet while the radioactive protein was in the solution.
• Conclusion: This experiment supported that T2 virus genes consist of DNA.

15.1.2 Molecular Properties of DNA

• Structure of a Deoxyribonucleotide:

  • Composed of a phosphate group, deoxyribose sugar, and a nitrogenous base (A, T, G, C).

• Strand Composition:

  • Strands consist of sugar-phosphate backbones with bases projecting outward.
  • Base Pairing Rules: A pairs with T, and G pairs with C held together by hydrogen bonds.

• Antiparallel Strands: 5' to 3' polarity running in opposite directions.

15.2 Semiconservative Replication Evidence: Meselson-Stahl Experiment

Experimental Setup

• E. coli Growth: Grown in $^{15}N$ (heavy nitrogen) and then transferred to $^{14}N$ (light nitrogen).
• Predictions:
  • If semiconservative, expect a mix of heavy and light densities after one division.
  • If conservative, first generation should show all heavy or light.
  • If dispersive, expect all hybrid densities.
• Results from Centrifugation:
  • First generation showed intermediate density (hybrid).
  • Second generation had lower density and some intermediate density.

Conclusion

• Data supports semiconservative replication since predictions held true for both generations.

15.3 Synthesis of Leading and Lagging Strands

15.3.1 Origin of Replication

• Bacterial Chromosome: Single origin of replication; DNA unwound at replication fork.
• Eukaryotic Chromosome: Multiple origins of replication to accommodate larger genome sizes.

15.3.2 Synthesis of Leading Strand

1. Primase synthesizes RNA primer for initiation.
2. DNA Polymerase synthesizes continuously in 5' to 3' direction as helicase unwinds DNA.

15.3.3 Synthesis of Lagging Strand

1. Okazaki Fragments: Synthesized discontinuously; each requiring a new RNA primer.
2. DNA polymerase replaces RNA primers with DNA.
3. DNA Ligase joins fragments to create a continuous strand.

15.4 Problems with Replicating Chromosome Ends

• Lagging Strand Issue: Final RNA primer removal leaves a single-strand end leading to chromosome shortening.
• Role of Telomerase: Extends unreplicated ends using its own RNA template, preventing loss of genetic information with cell divisions.

15.5 DNA Repair Mechanisms

Types of DNA Repair

• Direct Reversal: Simple fix for damaged DNA.
• Excision Repair: Removes damaged bases or nucleotides followed by DNA polymerase filling gaps.
• Mismatch Repair: Corrects DNA synthesis errors.
• Double-Strand Break Repair:
  • Homologous Recombination: Uses sister chromatid to repair.
  • Non-Homologous End Joining: Joins broken ends directly, with possible loss of some DNA.

Key Takeaways

• DNA Structure and Function: Underpins genetic inheritance and is crucial for accurate replication and repair.
• Repair Mechanisms: Essential for maintaining genetic integrity and preventing mutations that could lead to diseases like cancer.