Introduction. Chapter 11 DNA replication, repair and recombination. Overview. DNA replication is essential for life. Short on DNA structure

Transcription

1 Chapter 11 DNA replication, repair and recombination Overview Brief introduction DNA replication DNA repair DNA recombination DNA replication is essential for life Introduction Cells divide and make copies DNA must be copied DNA replication Must be accurate DNA repair Evolutionary change DNA recombination Short on DNA structure It has not escaped our notice that the specific basepairing we have postulated immediately suggests a possible copying mechanism for the genetic material - Watson and Crick, 1953 Basepairing: Adenine (A) binds thymine (T) Guanine (G) binds cytosine (C) 1

2 Basepairing enables DNA replication DNA replication Each strand a template for synthesizing a complementary strand DNA polymerase at the heart of replication Bacterial DNA Polymerases Synthesize DNA using a template Add nucleotides to 3 end of growing DNA chain Require: - DNA template - The four nucleoside triphosphates -Primer Li et al DNA Polymerase III Semiconservative replication - One new strand and one conserved strand 2

3 Replication begins at replication origins - Local areas where the stable DNA helix is opened up Multiple Origins in Eukaryotes Helicase breaks hydrogen bonds DNA polymerase Due to chromosome size and replication fork migration rate, eukaryotic chromosomes tend to have many origins for DNA replication Replication forks - Replication moves in both directions (bidirectional) The new strand is synthesized 5 3 Opposite directions DNA polymerase at work Results in problem at replication fork - Assymetrical with leading and lagging strand Leading and Lagging Strands Lagging strand Leading strand Discontinuous synthesis discovered by Okazaki Pulse-chase experiment revealed that new DNA was initially small pieces but became very large with time DNA polymerase synthesize 5 3 Problem at lagging strand! 3

4 DNA polymerase is self-correcting - Through error-correcting activity called proofreading -3 -to 5 exonuclease activity More important replication components - Topoisomerase to avoid tension build-up DNA DNA polymerase in work... Proofreading detects mispaired nucleotide and removes it (3 - to 5 ) Topoisomerase Correct nucleotide added and polymerase continues work... Helicase More important replication components - Single-strand binding protein prevents base pair re-forming - Sliding clamp keeps polymerase attached to DNA template Events at the replication fork - Summary Accuracy in replication important DNA repair Changes in DNA sequence often detrimental Sickle cell anemia as an example Permanent change mutation At the same time important for evolution! Need for DNA repair 4

5 DNA replication is very accurate DNA mismatch repair system - Corrects 99% of errors in the replication Examples of error rates US postal delivery of local first-class mail Airline luggage Professional typist Driving a car in the US DNA replication (no mismatch repair) DNA replication (with mismatch repair) 13 late deliveries per 100 parcels 1 lost bag per mistake per 250 characters 1 death per 10 4 people per year 1 mistake per 10 7 nucleotides 1 mistake per 10 9 nucleotides Hostile environment causes mutations - Such as thermal collisions, radiation and chemicals Purines A and G lost C converted to U Sources of Induced Mutations Fungal toxins Aflatoxin (peanuts) Cosmic rays UV light Chemical pollutants X rays Chemicals Industrial Cigarette smoke Hostile environment causes mutations - Such as thermal collisions, radiation and chemicals Basic mechanism of DNA repair 1. DNA damage removed by different nucleases 2. Repair DNA polymerase fills the gap 3. DNA ligase seals the nick 5

6 How can evolution act on DNA sequences? DNA replication and repair is VERY effective Maintains genetic message over millions of years Humans very similar to chimpanzees genetically ~98% similar...genetically! DNA recombination Genetic (DNA) recombination! Homologous recombination fundamental - Homology here referring to sequence similarity Mechanism of homologous recombination Regions with similar sequence Site-specific recombination - Between nonhomologous DNA sequences - Shuffle mobile genetic elements such as transposons - Very common ~45% of human genome Viruses are fully mobile genetic elements Mediates the movement of the transposon Sequences recognized by transposase Antibiotic resistance markers Extremely small Genes enclosed by a coat Contain RNA or DNA Single-stranded Double-stranded Herpes simplex virus 1 Ebola virus 6

7 Retroviruses common in Eucaryotic cells - DNA synthesized using RNA template (retro) - HIV is a retrovirus Reverse transcriptase 7