DNA Replication and Transcription: Biosynthesis of DNA and RNA 阮雪芬

DNA Replication and Transcription: Biosynthesis of DNA and RNA 阮雪芬 2004/05/21 @NTU

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

Introduction Replication Templates Repair

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

Characteristics of the Replication Process DNA Replication is Semiconservative Replication Begins at a Discrete Point on DNA

Two Possible Models for DNA Replication X

Density Gradient Centrifugation Matthew Meselson and Franklin Stahl 1957-1958

Preliminary Experiment

Actual Experiment

Replication Begins at a Discrete John Cairns, 1950 Point on DNA Replication fork Replication fork

The Theta Model for Replication of a Circular DNA Molecule In prokaryotes

Proposed Pathway fro Replication of Eukaryotic DNA

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

DNA Polymerase I Was isolated from E. coli, in 1957 dntp + (dnmp) n d(ntp ) n+1 + PPi

The Action of DNA Polymerase

Exonuclease Activity of DNA Acts as a proofreading and repair enzyme by catalyzing hydrolytic removal of mismatched bases. Polymerase I

5 3 Exonuclease Activity

Okazaki Fragments

Complete Scheme Showing Sequential Steps of Replication Helicase Process DNA gyrase: topoisomerase Formation of Replication fork

Single-stranded DNA Binding Proteins

Initiation of DNA Synthesis Primase

DNA Polymerase III

DNA Polymerase I

DNA Ligase

DNA Ligase

Eukaryotic Chromosomes and Telomeres Telomere: The repeating sequence AGGGTT Telomerase: catalyzes the synthesis of these DNA ends Telomerase becomes activated in human cancer cells

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

Mutations Spontaneous mutations Induced mutations

Spontaneous mutations Those changes that occur during normal genetic and metabolic functions in the cell. Two major types: Mismatching of base pairs: one for every 10 10 base pairs incorporated Base modifications caused by hydrolytic reactions

Spontaneous mutations Are favorable in that they confer a selective advantage to an organism. The causes of evolutionary processes Replication errors are of three types Substitution Insertion Deletion

The Basic Repair Mechanism for Mismatched Base Pairs in E. coli

Mutagenic Damage to DNA Caused by Spontaneous Processes DNA glycosidases

Induced mutations Mutagens Ionizing radiation Chemicals Heterocyclic base analogs Reactive chemical mutagens Intercalating agents

Ionizing radiation: Hydroxyl Radical

Chemical Mutagens Heterocyclic base analogs

Alkylating Agents

Alkylating Agents

Intercalating Agents

Binding of Intercalating Agents to Ames test DNA

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

Transcription

Initiation of Transcription

Elongation of the RNA Chain in Transcription

Termination of Transcription

Hairpin Loop

Inhibitors of Transcription

RNA-directed RNA Synthesis RNA replicase No editing, proofreading, or repair activities

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

Post-transcriptional Modification of trna in E. coli

mrna Processing Capping Poly A addition Splicing coding sequences

Capping Reactions

Poly A Addition 20 ~250 n Catalyzed by polyadenylate polymerase Increasing resistance to cellular nucleases

Splicing Coding Sequences

Outline Introduction Replication of DNA Action of DNA Polymerases DNA Damage and Repair Synthesis of RNA Post-transcriptional Modification of RNA Base Sequences in DNA

2, 3 -Dideoxyribonucleoside Triphosphates

Sanger Chain-termination Method

Exercises 11.1 a, c, d, f, k, l, m, n, s 11.2 11.8 11.9 11.10 11.15