BIOL10221 Molecular Biology DNA Replication I 2024 PDF

BIOL10221 Molecular Biology Module 2 - Lecture 1: DNA Replication I Ray O’Keefe [email protected] What will we cover? The possible different types of DNA replication Semiconservative replication and its experimental proof The role of topoisomerases in DNA replication...

BIOL10221 Molecular Biology Module 2 - Lecture 1: DNA Replication I Ray O’Keefe [email protected] What will we cover? The possible different types of DNA replication Semiconservative replication and its experimental proof The role of topoisomerases in DNA replication Why is it important? DNA needs to double to pass on equal amounts of DNA when cells divide DNA replication is therefore essential for reproduction and continued life Defects in DNA replication can cause diseases like cancer DNA topoisomerases solve the problem of DNA double helix unwinding caused by the separation of the two DNA strands during DNA replication Topoisomerases allow efficient and accurate replication of DNA BIOL10221 Module 2 - Lecture 1 2 DNA Replication The two DNA strands separate and DNA is copied using the parent strand as a template Complementary base pairing provides an obvious way for a double-stranded DNA molecule to replicate This is semiconservative replication End up with 2 identical strands same as parent DNA, each daughter strand has half the information from the parent BIOL10221 Module 2 - Lecture 3 1 Semiconservative replication is not the only possibility BIOL10221 Module 2 - Lecture 4 1 How might conservative replication work? BIOL10221 Module 2 - Lecture 5 1 How might dispersive replication work? In 1950’s this is what most scientists thought happened as of the three methods semiconservative was thought to be impossible! BIOL10221 Module 2 - Lecture 6 1 Why do we think about conservative and dispersive replication when the semiconservative mode looks so obvious? Because to begin with it was thought that semiconservative replication would be impossible Why? DNA is plectonemic – strands can’t be separated without unwinding the Double-helix structure Because many DNA molecules are circular (bacteria DNA, human mitochondria DNA, plasmids, DNA viruses) How do you unwind a circular molecule? BIOL10221 Module 2 - Lecture 7 1 Maybe you make a double-stranded cut, and then unwind it from the two ends linear, But still there are problems….. BIOL10221 Module 2 - Lecture 8 1 A typical bacterial chromosome is a circular DNA molecule of about 4,000,000 bp (4Mb) Imagine this is converted to a linear molecule A bacterium can divide every 20 minutes So it must replicate its genome every 20 minutes So it must replicate 200,000 bp per minute There are 10 bp per turn of the helix The DNA molecule would have to rotate at 20,000 revs per minute! BIOL10221 Module 2 - Lecture 9 1  The Meselson-Stahl experiment was carried out in 1959 It was designed to distinguish between the three modes of DNA replication and work out which occurs in E. coli bacteria (very simple and well designed experiment!) A culture of bacteria were grown in a medium containing heavy nitrogen The commonest isotope of nitrogen has an atomic weight of 14 – 14N The heavy isotope of nitrogen has an atomic weight of 15 – 15N Nitrogen component of the DNA bases so can get into DNA BIOL10221 Module 2 - Lecture 10 1 The culture medium contained 15NH4Cl - heavy ammonium chloride All of the bacteria’s DNA became labelled with 15N N-DNA can be distinguished from 14N-DNA by 15 density gradient centrifugation Buoyant density CsCl = cesium chloride BIOL10221 Module 2 - Lecture 11 1 The Meselson-Stahl experiment BIOL10221 Module 2 - Lecture 12 1 BIOL10221 Module 2 - Lecture 13 1 The Meselson-Stahl experiment showed that in living cells, DNA is replicated by the semiconservative process So how are the strands separated without unwinding the DNA double helix? This is the job of enzymes called DNA topoisomerases BIOL10221 Module 2 - Lecture 14 1 DNA Supercoils are produced if strands are separated These supercoils need to be removed or prevented DNA topoisomerases solve this problem BIOL10221 Module 2 - Lecture 15 1 The mode of action of a Type I DNA topoisomerase BIOL10221 Module 2 - Lecture 16 1 How a Type I DNA topoisomerase separates the strands without helix unwinding during DNA replication BIOL10221 Module 2 - Lecture 17 1 The mode of action of a Type II DNA topoisomerase (major enzyme used) BIOL10221 Module 2 - Lecture 18 1  Topoisomerase type II mechanism G Segment – DNA strand that gets completely cut so other strand can pass through T Segment – DNA strand that gets passed through BIOL10221 Module 2 - Lecture 19 1 https://youtu.be/k4fbPUGKurI BIOL10221 Module 2 - Lecture 20 1 Resources Reading BROWN, Chapter 10 pages 187-191, 192-193 ALBERTS, Chapter 6 pages 199-205, 212 Blackboard PDF files with Objectives, Terms, Questions and Answers BIOL10221 Module 2 - Lecture 21 1  BIOL10221 Molecular Biology Module 2 - Lecture 1: DNA Replication I Ray O’Keefe [email protected]