Biological Science - Chapter 15: DNA and the Gene: Synthesis & Repair

Third Canadian Edition Biological Science

Overview

  • Authors: Freeman, Quillin, Allison, Black, Podgorski, Taylor Carmichael, Harrington, Sharp.

  • Course: BLG143: Biology I

  • Chapter Focus: DNA and the Gene: Synthesis & Repair

  • Affiliation: Department of Chemistry & Biology, Toronto Metropolitan University.

What Are Genes Made Of?

  • Chromosomes: Comprised of DNA and protein.

  • Controversy: The debate on whether genes are made of DNA or protein.

    • Initial thought: Most biologists believed genes were proteins due to their complexity and variability.

    • Reality: DNA consists of only four different nucleotides.

The Hershey–Chase Experiment

  • Study Focus: How the T2 virus infects Escherichia coli.

    • Process: Virus injects genes into the bacterium, directing new virus production while leaving its protein coat (capsid) outside.

  • Key Question: Does protein or DNA enter the E. coli cell?

  • Experiment Details:

    • Growth of virus with radioactive isotopes:

    • 32P: Incorporated into DNA.

    • 35S: Incorporated into proteins.

    • Result: Only radioactive DNA found inside cells, confirming genes must be DNA.

The Secondary Structure of DNA

  • Primary Structure Components:

    1. Backbone: Sugar and phosphate groups of deoxyribonucleotides.

    2. Nitrogen-Containing Bases: Project from the backbone.

  • Directionality: 3′ end: exposed hydroxyl group; 5′ end: exposed phosphate group.

  • Nucleobases:

    • Purines (double ring): Adenine (A), Guanine (G).

    • Pyrimidines (single ring): Thymine (T), Cytosine (C), Uracil (found in RNA).

    • Complementary pairs: A+T and G+C.

Watson and Crick Model of DNA

  • Antiparallel Strands: Two DNA strands aligned in opposite directions.

  • Double Helix Formation: The structure of DNA stabilized by complementary base pairing.

  • Hydrogen Bonding Patterns:

    • A pairs with T (2 hydrogen bonds).

    • G pairs with C (3 hydrogen bonds).

DNA Replication Hypotheses

  1. Semiconservative Replication: Each daughter DNA contains one parental and one new strand.

  2. Conservative Replication: An entirely new molecule is formed alongside the original; each daughter would then have either all old or all new strands.

  3. Dispersive Replication: Parent DNA is fragmented, resulting in daughters with interspersed old and new DNA.

The Meselson-Stahl Experiment

  • Methodology:

    • E. coli grown in 15N (heavy nitrogen) first.

    • Transferred to 14N (normal nitrogen).

    • DNA collected after cell divisions, separated by density through centrifugation.

  • Results: Supported the semiconservative replication model; data rejected conservative and dispersive hypotheses based on banding patterns.

Mechanics of DNA Synthesis

Enzymatic Functions in Replication

  • DNA Polymerase: Enzyme that catalyzes DNA synthesis, working in one direction only (5′→3′).

  • Deoxyribonucleoside Triphosphates (dNTPs): Precursors for DNA synthesis, carrying high potential energy due to packed phosphate groups.

  • Replication Origin:

    • Formation of replication bubbles at specific sequences.

    • Bacteria: One origin causing one bubble.

    • Eukaryotes: Multiple origins per chromosome, each with two replication forks for bidirectional synthesis.

Unwinding and Stabilizing DNA

  • Helicase: Unwinds the double helix by breaking hydrogen bonds.

  • Single-Strand Binding Proteins (SSBPs): Prevent re-annealing of single strands post-unwinding.

  • Topoisomerase: Relieves tension by cutting and rejoining DNA strands.

Leading and Lagging Strand Synthesis

  • Leading Strand: Synthesized continuously toward the replication fork (5′ to 3′) using primers provided by the enzyme primase (an RNA polymerase).

  • Lagging Strand: Synthesized discontinuously away from the fork, producing Okazaki fragments linked by DNA ligase after primer removal.

  • Okazaki Fragments: Short DNA pieces on the lagging strand formed between sections of RNA primer.

Telomeres and Chromosome Replication Problems

  • Lagging Strand Issues: Replication cannot fully replicate the ends due to the absence of a primer.

  • End Replication Problem: Results in single-stranded DNA at lagging strand ends, leading to chromosome shortening over time.

  • Telomere Function: Composed of repetitive non-coding DNA, protecting important gene sequences from degradation.

Telomerase Enzyme

  • Function: Catalyzes the synthesis of telomere DNA using its own RNA template, preventing the shortening of telomeres during replication.

  • Discovery: Elizabeth Blackburn's research on Tetrahymena cells led to the identification of telomerase.

DNA Repair Mechanisms

  • DNA Replication Accuracy:

    • Error rate: 1 mistake per billion bases with proofreading enzymes.

  • DNA Polymerase Proofreading:

    • Involves the removal of mismatched bases, improving fidelity.

  • Mismatch Repair System: Post-replication correction of mismatches including base removal and correct base insertion.

  • Nucleotide Excision Repair: Repairs damage caused by chemical and physical agents by excising damaged sections and synthesizing replacements.

Implications of Telomeres in Aging and Cancer

  • Somatic Cell Behavior: Lack telomerase, leading to chromosome shortening as cells divide, implicating aging.

  • Cancer Cell Mechanisms: Often maintain telomere length through continuous telomerase activity, allowing indefinite division.

  • Research on Therapies: Potential treatments targeting telomerase activity to control cancer cellular proliferation have yielded varied responses in experimental settings.

Summary and Learning Objectives

  • Primary focus on DNA structure, synthesis, replication mechanisms, and their relation to heredity and cellular processes.

  • Understanding of telomeres, their biological significance, and relevance to aging and cancer treatment is emphasized.