DNA: The Molecular Basis of Inheritance – Study Notes 16

DNA: The Molecular Basis of Inheritance

Key Learning Objectives

• Describe DNA structure & evidence that DNA is the genetic material.

• Compare prokaryotic vs. eukaryotic DNA replication.

• Explain the Meselson-Stahl experiment and semiconservative replication.

• Identify chromatin packaging features.

• Understand enzymes of DNA replication.

• Explain DNA repair mechanisms.

• Understand end replication problem in eukaryotes.

Evidence for DNA as Genetic Material

Seminal Experiments

• Frederick Griffith (1928): Discovered transformation using S and R strains of Streptococcus pneumoniae.

• Avery, MacLeod, McCarty (1944): Identified DNA as the transforming substance.

• Hershey & Chase (1952): Used bacteriophage T2 to show DNA, not protein, is the genetic material.

• Chargaff's Rules: Amount of A = T and G = C in DNA.

• Watson & Crick (1953): Proposed the double helix structure of DNA based on Rosalind Franklin's X-ray diffraction data.

Structure of DNA

• Double helix: Two antiparallel strands.

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

• Base pairing: A pairs with T (2 hydrogen bonds), G pairs with C (3 hydrogen bonds).

• Phosphodiester bonds: Link nucleotides in a strand.

• Antiparallel orientation: One strand runs 5' to 3', the other 3' to 5'.

DNA Replication Basics

• Semiconservative replication: Each new DNA molecule consists of one old and one new strand.

• Meselson-Stahl Experiment: Used isotopic labeling to demonstrate semiconservative replication.

Origins of Replication

• Prokaryotes: Single origin, circular DNA.

• Eukaryotes: Multiple origins, linear chromosomes.

Enzymes of DNA Replication Fork

• Helicase: Unwinds DNA double helix.

• Single-Strand Binding Proteins (SSB): Stabilize unwound DNA.

• Topoisomerase: Relieves supercoiling ahead of the fork.

• Primase: Synthesizes short RNA primers.

• DNA Polymerase III: Synthesizes new DNA strand by adding nucleotides to the primer.

• DNA Polymerase I: Replaces RNA primers with DNA.

• Ligase: Joins Okazaki fragments on the lagging strand.

Leading vs. Lagging Strand Synthesis

• Leading strand: Synthesized continuously in the 5' to 3' direction.

• Lagging strand: Synthesized discontinuously as Okazaki fragments.

• Okazaki fragments: Short DNA segments on the lagging strand, later joined by ligase.

DNA Repair Mechanisms

• Proofreading: DNA polymerases correct errors during replication.

• Mismatch repair: Enzymes remove and replace incorrectly paired nucleotides.

• Nucleotide excision repair: Removes bulky lesions (e.g., thymine dimers) using nucleases.

Chromatin Structure

• Euchromatin: Loosely packed, transcriptionally active DNA.

• Heterochromatin: Densely packed, transcriptionally inactive (centromeres, telomeres).

End Replication Problem

• Eukaryotic chromosomes: Shorten with each replication due to incomplete replication of 5' ends.

• Telomerase: Enzyme that extends telomeres in germ cells and some stem cells.

• Prokaryotes: Circular DNA, so end replication problem does not occur.

Summary Table: Key Enzymes in DNA Replication

Enzyme	Function
Helicase	Unwinds DNA double helix
SSB Proteins	Stabilize single-stranded DNA
Topoisomerase	Relieves supercoiling
Primase	Synthesizes RNA primers
DNA Polymerase III	Main DNA synthesis
DNA Polymerase I	Replaces RNA primers with DNA
Ligase	Joins Okazaki fragments

Key Equations

• Base pairing:

$\text{A} = \text{T}$ $\text{G} = \text{C}$

• Direction of DNA synthesis:

$\text{DNA is synthesized in the 5' \rightarrow 3' direction}$

Example: Meselson-Stahl Experiment

Bacteria were grown in heavy ($^{15}$N) and light ($^{14}$N) nitrogen media. After one round of replication in light media, DNA had intermediate density, supporting semiconservative replication.

Additional info: Chromatin structure and DNA repair mechanisms are essential for maintaining genome stability and proper gene expression.