Lecture Molecular biology (Fifth Edition): Chapter 2 - Robert F. Weaver

Before we begin to study in detail the structure and activities of genes, and the experimental evidence underlying those concepts, we need a fuller outline of the adventure that lies before us. Thus, in this chapter and in chapter 3, we will fl esh out the brief history of molecular biology presented in chapter 1. In this chapter we will begin this task by considering the behavior of genes as molecules.
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Lecture Molecular biology (Fifth Edition): Chapter 2 - Robert F. WeaverLecture PowerPoint to accompanyMolecular Biology Fifth Edition Robert F. Weaver Chapter 2The Molecular Nature of Genes Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display. The Nature of Genetic MaterialHistorical Background• Miescher isolated nuclei from pus (white blood cells) in 1869 – Found a novel phosphorus-bearing substance = nuclein – Nuclein is mostly chromatin, a complex of DNA and chromosomal proteins• End of 19th century – DNA and RNA separated from proteins• Levene, Jacobs, et al. characterized basic composition of DNA and RNA 2-2Molecular Foundation: Early experiments that explored the question: What is the genetic material?• Key experiments performed by Frederick Griffith in 1928• Observed change in Streptococcus pneumoniae — from avirulent (R) rough colonies, bacteria without capsules, to virulent (S) smooth colonies, bacteria that had capsules• Result: Heat-killed virulent bacteria could transform avirulent bacteria into virulent bacteria 2-3Outline of Griffith’s Transformation Experiments 2-4 DNA: The Transforming MaterialIn 1944 Avery, Macleod and McCarty used atransformation test similar to Griffith’sprocedure taking care to define the chemicalnature of the transforming substance – Techniques used excluded both protein and RNA as the chemical agent of transformation – Exclusion of DNA verified that DNA is the chemical agent of transformation of S. pneumoniae from avirulent to virulent 2-5 Analytical ToolsPhysical-chemical analysis has often used:1. Ultracentrifugation Used to estimate size of material1. Electrophoresis Indicated high charge-to-mass ratio1. Ultraviolet Absorption Spectrophotometry Absorbance of UV light matched that of DNA1. Elementary Chemical Analysis Nitrogen-to-phosphorus ratio of 1.67, expected for DNA but lower than expected for protein 2-6Confirmation for DNA as the genetic material• In the 1940s geneticists doubted the use of DNA as the genetic material as it appeared to be monotonous repeats of 4 bases• By 1953 Watson & Crick published the double- helical model of DNA structure and Chargaff demonstrated that the 4 bases were not present in equal proportions• In 1952 Hershey and Chase demonstrated that bacteriophage infection comes from DNA, adding more evidence to support that DNA is the genetic material 2-7Outline of Hershey and Chase’s Experiment 2-8 Summary• The classic molecular biology experiments performed by Griffith, Avery, MacLeod, Mccarty, Hershey and Chase combined revealed that DNA is the genetic element 2-9 The Chemical Nature of Polynucleotides• Biochemists determined the components of nucleotides during the 1940s• The component parts of DNA – Nitrogenous bases: • Adenine (A) • Cytosine (C) • Guanine (G) • Thymine (T) – Phosphoric acid – Deoxyribose sugar 2-10 Nucleosides and Deoxyribose• RNA component parts – Nitrogenous bases • Like DNA except Uracil (U) replaces Thymine – Phosphoric acid – Ribose sugar• Bases use ordinary numbers• Carbons in sugars are noted as primed numbers• Nucleotides contain phosphoric acid • Deoxyribose lacks a• Nucleosides lack the hydroxyl group (OH) at phosphoric acid the 2-position 2-11 Purines and Pyrimidines• Adenine and guanine are related structurally to the parent molecule purine• Cytosine, thymine and uracil resemble the parent molecule pyrimidine 2-12 DNA Linkage• Nucleotides are nucleosides with a phosphate group attached through a phosphodiester bond• Nucleotides may contain one, two, or even three phosphate groups linked in a chain 2-13 A TrinucleotideThe example trinucleotidehas polarity – The top of molecule has a free 5’-phosphate group = 5’ end – The bottom has a free 3’- hydroxyl group = 3’ end 2-14 Summary• DNA and RNA are chain-like molecules composed of subunits called nucleotides• Nucleotides contain a base linked to the 1’-position of a sugar and a phosphate group• The phosphate joins the sugars in a DNA or RNA chain through their 5’- and 3’- hydroxyl groups by phosphodiester bonds ...