DNA and the Gene: Synthesis and Repair – Study Notes

DNA and the Gene: Synthesis and Repair

Introduction

This chapter explores the molecular nature of genes, focusing on the structure of DNA, the mechanisms of DNA replication, and the processes involved in repairing DNA damage. Understanding these concepts is fundamental to genetics and cell biology.

What Are Genes Made Of?

DNA vs. Protein as Genetic Material

• Chromosomes are composed of DNA and proteins.

• Historically, proteins were thought to be the genetic material due to their complexity and variability.

• DNA is made of only four nucleotides, leading to initial skepticism about its role as genetic material.

The Hershey–Chase Experiment

• Studied T2 virus infection in Escherichia coli.

• Used radioactive isotopes to label DNA (phosphorus) and protein (sulfur).

• Only radioactive DNA entered bacterial cells, demonstrating that DNA is the hereditary material.

The Structure of DNA

DNA as a Double Helix

• DNA is a double-stranded polymer of deoxyribonucleotides.

• Each nucleotide consists of a deoxyribose sugar, a phosphate group, and a nitrogenous base.

• Nucleotides are linked by phosphodiester bonds between the 3′ hydroxyl and 5′ phosphate groups.

Directionality and Antiparallel Strands

• Each DNA strand has a 5′ end (phosphate group) and a 3′ end (hydroxyl group).

• Strands are antiparallel and twist to form a double helix.

• Complementary base pairing: Adenine (A) pairs with Thymine (T); Guanine (G) pairs with Cytosine (C).

• The double helix is stabilized by hydrogen bonds and base stacking interactions.

Testing Early Hypotheses About DNA Synthesis

Models of DNA Replication

• Semiconservative replication: Each daughter DNA has one old and one new strand.

• Conservative replication: One daughter has both old strands, the other both new.

• Dispersive replication: Each daughter has interspersed old and new DNA segments.

The Meselson–Stahl Experiment

• Used isotopes of nitrogen to label DNA in E. coli.

• Demonstrated that DNA replication is semiconservative.

A Model for DNA Synthesis

DNA Polymerase and Directionality

• DNA polymerase catalyzes DNA synthesis, adding nucleotides only to the 3′ end.

• DNA synthesis proceeds in the 5′ → 3′ direction.

• Monomers are deoxyribonucleoside triphosphates (dNTPs), which provide energy for bond formation.

Origin of Replication and Replication Forks

• Replication begins at specific sequences called origins of replication.

• Bacteria have one origin; eukaryotes have many per chromosome.

• Replication bubbles form, each with two replication forks moving bidirectionally.

Opening and Stabilizing the Helix

• DNA helicase separates DNA strands by breaking hydrogen bonds.

• Single-strand DNA-binding proteins (SSBPs) prevent re-annealing.

• Topoisomerase relieves tension ahead of the replication fork by cutting and rejoining DNA.

Synthesis of Leading and Lagging Strands

Leading Strand Synthesis

• DNA polymerase requires a primer (short RNA strand) to begin synthesis.

• Primase synthesizes the RNA primer.

• The leading strand is synthesized continuously toward the replication fork in the 5′ → 3′ direction.

Lagging Strand Synthesis and Okazaki Fragments

• The lagging strand is synthesized discontinuously, away from the replication fork.

• Primase synthesizes multiple RNA primers as the fork opens.

• DNA polymerase synthesizes short DNA fragments (Okazaki fragments).

• DNA polymerase I removes RNA primers and replaces them with DNA.

• DNA ligase joins Okazaki fragments by forming phosphodiester bonds.

Protein	Function
DNA polymerase III	Main enzyme for DNA synthesis
Primase	Synthesizes RNA primers
DNA ligase	Joins Okazaki fragments
Helicase	Unwinds DNA helix
SSBPs	Stabilize single strands
Topoisomerase	Relieves supercoiling
DNA polymerase I	Removes RNA primers, fills in DNA

Replicating the Ends of Linear Chromosomes

Telomeres and the End Replication Problem

• Telomeres are repetitive DNA sequences at chromosome ends.

• Lagging strand synthesis leaves a single-stranded overhang, leading to chromosome shortening with each division.

• Telomeres do not encode genes but protect chromosome integrity.

Role of Telomerase

• Telomerase extends the 3′ end using its own RNA template, allowing complete replication of chromosome ends.

• Active mainly in gametes and stem cells; most somatic cells lack telomerase, leading to progressive shortening.

• Telomerase activity is associated with cancer cell immortality.

Repairing Mistakes and DNA Damage

Accuracy of DNA Replication

• DNA polymerase has high fidelity, with an error rate of about one mistake per billion nucleotides.

• Proofreading by DNA polymerase corrects most errors during synthesis.

Mismatch Repair

• Mismatch repair enzymes correct errors missed by DNA polymerase after replication.

• They recognize, remove, and replace incorrect bases in the newly synthesized strand.

Repairing Damaged DNA

• DNA can be damaged by UV light, X-rays, and chemicals (e.g., thymine dimers).

• Nucleotide excision repair removes damaged DNA and fills in the correct sequence using the undamaged strand as a template.

• DNA ligase seals the repaired strand.

DNA Repair Defects and Disease

• Xeroderma pigmentosum (XP) is a rare genetic disorder caused by mutations in nucleotide excision repair genes.

• XP results in extreme sensitivity to UV light and a high risk of skin cancer.

• Defects in DNA repair genes are often linked to cancer due to increased mutation rates.