Week 8 - Oct 31

DNA Structure and Replication

Introduction to DNA Structure

Deoxyribonucleic acid (DNA) is the hereditary material in all living organisms. Its double helix structure allows for the accurate storage and transmission of genetic information.

• Double Helix: DNA consists of two antiparallel strands twisted into a double helix, with complementary base pairing (A-T, G-C).

• Nucleotides: Each strand is composed of nucleotides, each containing a phosphate group, deoxyribose sugar, and a nitrogenous base.

• Base Pairing: Adenine pairs with thymine via two hydrogen bonds; guanine pairs with cytosine via three hydrogen bonds.

Replication Errors and DNA Damage

Sources and Consequences of DNA Damage

DNA can be damaged by various internal and external factors, leading to mutations if not properly repaired.

• Replication Errors: Mistakes during DNA synthesis can introduce incorrect bases.

• Chemical Agents: Both natural and synthetic chemicals can alter DNA structure.

• Physical Agents: Radioactivity, X-rays, and ultraviolet (UV) radiation can damage DNA. For example, UV light can cause thymine dimers, where adjacent thymine bases bond together, distorting the DNA helix.

• Strand Damage: Damage can affect one or both DNA strands, potentially leading to mutations if unrepaired.

DNA Proofreading and Repair during Replication

DNA Polymerase Proofreading

DNA polymerase has a proofreading function that corrects errors during DNA synthesis, ensuring high fidelity in DNA replication.

• Detection of Errors: Shape distortions caused by mismatched bases do not fit properly in the polymerase active site, causing polymerization to pause.

• Exonuclease Activity: The 3' end of the new DNA strand is displaced to the exonuclease site of the polymerase, where the incorrect nucleotide is removed.

• Resumption of Synthesis: DNA synthesis resumes after the error is excised and the correct nucleotide is added.

Equation:

$\text{DNA}_{(n)} + dNTP \xrightarrow{\text{DNA polymerase}} \text{DNA}_{(n+1)} + PP_i$

Repair Continues after DNA Replication

Mismatch Repair Mechanism

Mismatch repair corrects errors that escape proofreading, acting immediately after DNA synthesis.

• Recognition: Mismatch repair enzymes detect mismatched bases in the newly synthesized DNA strand.

• Excision: The section of the new strand containing the incorrect base is removed.

• Resynthesis: DNA polymerase fills in the correct bases.

• Ligation: DNA ligase seals the nicks in the sugar-phosphate backbone.

Fidelity: After proofreading and mismatch repair, the error rate is about one mistake per billion base pairs ($1 \times 10^9$).

Different DNA Repair Mechanisms

Nucleotide Excision Repair (NER)

NER is a versatile repair system that removes bulky DNA lesions, such as thymine dimers caused by UV light.

• Damage Recognition: Repair proteins scan DNA to identify specific base alterations.

• Excision: Enzymes cut the DNA on both sides of the lesion, excising the damaged segment.

• Resynthesis: DNA polymerase fills in the gap with new nucleotides.

• Ligation: DNA ligase joins the newly synthesized segment to the rest of the strand.

Summary Table: Steps in Nucleotide Excision Repair

Step	Description
1. Error Detection	Repair proteins recognize DNA damage
2. DNA Unwinding	Helicase unwinds the DNA around the lesion
3. Nucleotide Excision	Endonucleases cut on both sides of the lesion, removing the damaged DNA
4. Repair Synthesis	DNA polymerase synthesizes new DNA to fill the gap
5. Ligation	DNA ligase seals the nick in the sugar-phosphate backbone

DNA Replication Overview

Key Steps and Enzymes in DNA Replication

DNA replication is a highly coordinated process involving several enzymes and steps to ensure accurate duplication of genetic material.

• 1. Opening the Helix:

  • Helicase: Catalyzes the breaking of hydrogen bonds to separate DNA strands.

  • Single-strand DNA-binding proteins (SSBPs): Stabilize single-stranded DNA.

  • Topoisomerase: Relieves tension caused by unwinding the DNA helix.

• 2. Leading Strand Synthesis:

  • Primase: Synthesizes RNA primer to provide a starting point for DNA synthesis.

  • DNA polymerase III: Extends the leading strand continuously.

• 3. Lagging Strand Synthesis:

  • Primase: Synthesizes RNA primers for Okazaki fragments.

  • DNA polymerase III: Synthesizes Okazaki fragments discontinuously.

  • DNA polymerase I: Removes RNA primers and replaces them with DNA.

  • DNA ligase: Joins Okazaki fragments to form a continuous strand.

Summary Table: Major Enzymes in DNA Replication

Enzyme	Function
Helicase	Unwinds the DNA double helix
SSBPs	Stabilize single-stranded DNA
Topoisomerase	Relieves supercoiling ahead of the replication fork
Primase	Synthesizes RNA primers
DNA polymerase III	Extends DNA from the primer
DNA polymerase I	Removes RNA primers and fills in with DNA
DNA ligase	Joins DNA fragments together

Example: During exposure to UV light, thymine dimers may form. Nucleotide excision repair removes these dimers, preventing mutations that could lead to diseases such as skin cancer.

Additional info: The high fidelity of DNA replication is essential for maintaining genetic stability across generations. Defects in DNA repair mechanisms can lead to genetic disorders and increase susceptibility to cancer.