Molecular Basis of Inheritance

Molecular Basis of Inheritance

Introduction

  • Presenter: Bethanie-Michelle Statler

Griffith Experiments

  • Key Discoveries: Frederick Griffith discovered that a molecule within cells provides genetic material.

  • Types of Bacteria:

    • Pathogenic bacteria (S cells), capable of causing disease.

    • Harmless bacteria (R cells), non-pathogenic.

  • Experimental Results: Mixing heat-killed S cells with R cells resulted in the death of mice.

  • Transformation: The process where a cell incorporates foreign DNA into its genome, leading to genetic changes.

Viral DNA and Transforming Substance

  • Key Contributors: Oswald Avery, Maclyn McCarty, and Colin MacLeod identified the transforming substance as DNA.

  • Bacteriophages (Phages): Viruses that specifically infect bacteria, composed of either DNA or RNA enclosed by a protein coat.

DNA as Genetic Material

  • 1952 Experiment by Alfred Hershey and Martha Chase: Demonstrated that DNA is the genetic material of phage T2.

  • Experiment Details - Batch 1:

    • Labeled phages with radioactive sulfur ($^{35}S$) in phage protein.

    • Phages infect cells; agitation frees outside phage parts from cells.

    • Cells subjected to centrifugation; radioactivity (phage protein) found in liquid.

  • Experiment Details - Batch 2:

    • Labeled phages with radioactive phosphorus ($^{32}P$) in phage DNA.

    • Similar procedures followed, resulting in radioactivity (phage DNA) being found in the pellet.

Structure of DNA

  • Composition of DNA: DNA is a polymer of nucleotides, each nucleotide consists of:

    • A nitrogenous base

    • A sugar

    • A phosphate group

  • Nitrogenous Bases:

    • Adenine (A)

    • Thymine (T)

    • Guanine (G)

    • Cytosine (C)

  • Chargaff’s Rules (1950):

    • DNA composition varies among species.

    • In any species, the quantity of A equals T (A=T), and quantity of G equals C (G=C).

DNA Double-Strand Structure

  • Covalent Bonds: Nucleotides linked via covalent phosphodiester bonds in one strand.

  • Hydrogen Bonds: Nitrogenous bases form hydrogen bonds with a complementary second strand.

  • Antiparallel Nature: DNA strands run in opposite directions (3' to 5' and 5' to 3').

X-ray Crystallography and DNA Discoveries

  • Contributors: Maurice Wilkins and Rosalind Franklin employed X-ray crystallography to study DNA.

  • Franklin's Contribution: Produced a critical picture of DNA, aiding in structural understanding.

  • Watson and Crick’s Model: Deduced the structure of DNA as a double helix using Franklin's data. Recognized for their contribution with the Nobel Prize in 1962.

Key Features of DNA Structure

  • Dimensions:

    • Distance between base pairs: 0.34 nm

    • Width of DNA double helix: 2 nm

    • Complete turn of the helix: 3.4 nm

  • Base Pairing:

    • A pairs with T

    • G pairs with C

DNA Replication - Semiconservative Model

  • Replication Process: Watson and Crick’s model predicts each daughter molecule consists of one old strand and one new strand.

  • Competing Models:

    • Conservative model: Parent strands rejoin post-replication.

    • Dispersive model: Each strand is a mix of old and new DNA.

  • Experimental Support: Matthew Meselson and Franklin Stahl's experiments supported the semiconservative model.

Mechanism of DNA Replication

  • Origins of Replication: Sites where DNA strands are separated, forming a replication “bubble”.

  • Directionality of Replication: Replication occurs bidirectionally from each origin until the entire molecule is copied.

  • Replication Fork: The Y-shaped region where new DNA strands elongate.

  • Key Enzymes:

    • Helicases: Untwist the DNA double helix.

    • Single-strand Binding Proteins: Stabilize single-stranded DNA.

    • Topoisomerases: Correct the overwinding ahead of replication forks by breaking, swiveling, and rejoining DNA strands.

DNA Polymerization Process

  • Synthesis Direction: DNA polymerase can only synthesize DNA in the 5' to 3' direction.

  • Role of Primase: Synthesizes a small RNA primer to initiate DNA synthesis, generally 5-10 nucleotides long.

  • DNA Polymerase Activity: DNA polymerase (DNA pol III) begins elongating DNA by adding nucleotides to the 3' end of an RNA primer. DNA pol I replaces RNA primers with DNA.

Antiparallel Elongation Details

  • Leading Strand: Synthesized continuously towards the replication fork.

  • Lagging Strand: Synthesized discontinuously in fragments called Okazaki fragments, starting closest to the origin of replication.

  • Joining of Fragments: DNA ligase joins Okazaki fragments together after synthesis.

DNA Replication Checkpoint Questions

  1. Untwisting the Double Helix: Which enzyme? (Helicase)

  2. Adding Nucleotides: Which enzyme adds DNA nucleotides to the 3’ end? (DNA pol III)

  3. Preventing Overwinding: Which enzyme prevents overwinding of the DNA double helix? (Topoisomerase)

  4. Adding RNA Primers: Which enzyme adds an RNA primer? (Primase)

DNA Proofreading and Repair Mechanisms

  • Proofreading by DNA Polymerases: Corrects errors by replacing incorrect nucleotides during DNA synthesis.

  • Mismatch Repair: Repair enzymes replace incorrectly paired nucleotides that evade proofreading mechanisms.

  • Normal Mutation Rates: A low rate of mutation contributes to genetic variation.

  • Causes of DNA Damage: Harmful chemical agents (e.g., cigarette smoke) and physical agents (e.g., X-rays).

  • Nucleotide Excision Repair: A nuclease removes damaged DNA sections and replaces them with proper nucleotides.

Challenges with Ends of Linear DNA

  • Limitations of Replication Machinery: Cannot complete the 5' ends of daughter DNA strands, leading to progressively shorter strands.

  • Prokaryotic vs Eukaryotic DNA: Circular chromosomes in prokaryotes avoid this issue.

Telomeres in Eukaryotic Cells

  • Function of Telomeres: Special nucleotide sequences at the ends of eukaryotic chromosomes.

  • Conservation of Genes: Telomeres do not prevent DNA shortening but postpone erosion of essential genes.

  • Telomerase Activity: Enzyme found in germ cells (egg and sperm) catalyzing telomere lengthening. Shortening of telomeres may help prevent cancer by limiting cell division.

  • Cancer Cells: Some exhibit telomerase activity, which causes continuous division and survival.

Chromosomal Structure and Organization

  • Bacterial Chromosomes: Double-stranded, circular DNA associated with minimal protein, supercoiled within a nucleoid region of the cell.

  • Eukaryotic Chromosomes: Contain linear DNA associated with extensive proteins; the DNA forms chromatin that is compacted to fit into the nucleus.

  • Histones: Proteins essential for chromatin folding and organization.

Nucleosome and DNA Packing

  • Nucleosome Structure: DNA wound around a core of eight histones, forming a 10-nm fiber.

  • Chromatin Types:

    • Euchromatin: Loosely packed, accessible for transcription.

    • Heterochromatin: Highly condensed, transcriptionally inactive regions.

  • Chromatin Organization:

    • 30-nm fiber: Compacted structure formed by histones.

    • Looped domains: Further compaction, forming metaphase chromosomes, enabling the orderly segregation of genetic material during cell division.