Biophysics

Preface vii 1. Laws of Physics and Chemistry 1 1.1 1.2 1.3 1.4 1.5 Introduction 1 Quantum Mechanics 1 The Electronic Structure of Atoms 3 Molecular Orbitals andCovalent Bonds 5 Molecular Interactions 8 1.5.1 1.5.2 Strong interactions 9 Weak interactions 10 1.6 Stereochemistry andC hirality 11 1.6.1 Stereochemical nomenclature 13 1.7 Thermodynamics 13 1.7.1 1.7.2 1.7.3 1.7.4 1.7.5 Entropy 16 Enthalpy 17 The free energy of a system 17 Chemical potential 18 Oxidation-reduction potential 19 1.8 Radioactivity 20 1.8.1 1.8.2 1.8.3 1.8.4 1.8.5 Rate of radioactive decay 21 Measurement of radioactivity 22 Effects of radioactivity on matter 22 Biological effects of radiation 23 Applications of radio isotopes 23 2. Separation Techniques 24 2.1 2.2 Introduction 24 Chromatography 24 2.2.1 Column chromatography 26 x Contents 2.2.2 2.2.3 2.2.4 2.2.5 2.2.6 2.2.7 2.2.8 2.2.9 Thin layer chromatography 26 Paper chromatography 26 Adsorption chromatography 27 Partition chromatography 27 Gas liquid chromatography (GLC) 28 Ion exchange chromatography 28 Molecular exclusion chromatography 29 Affinity chromatography 3 0 2.3 Electrophoresis 3 2 2.3.1 2.3.2 2.3.3 2.3.4 2.3.5 2.3.6 2.3.7 2.3.8 Moving boundary electrophoresis 3 2 Zone electrophoresis 3 3 Low voltage electrophoresis 3 3 High voltage electrophoresis 3 3 Gel electrophoresis 3 4 Sodium dodecyl sulphate poly acrylamide gel electrophoresis (SDS-PAGE) Iso electric focussing 3 5 Continuous flow electrophoresis 3 5 3. Physico-Chemical Techniques to Study Biomolecules 37 3.1 3.2 3.3 3.4 3.5 3.6 3.7 3.8 Introduction 3 7 Hydration of Macromolecules 3 8 Role of Friction 3 8 Diffusion 3 9 Sedimentation 41 The Ultracentrifuge 43 Viscosity 46 Rotational Diffusion 48 3.8.1 3.8.2 Flow birefringence measurements 49 Electric birefringence 50 3.9 Light Scattering 51 3.10 Small Angle X-ray Scattering 54 4. Spectroscopy 58 4.1 4.2 4.3 4.4 4.5 4.6 4.7 Introduction 58 Ultraviolet/Visible Spectroscopy 59 Circular Dichroism (CD) and Optical Rotatory Dispersion (ORD) 61 Fluorescence Spectroscopy 65 Infrared Spectroscopy 67 Raman Spectroscopy 71 Electron Spin Resonance 74 5. Light Microscopy 77 5.1 5.2 5.3 5.4 Introduction 77 Elementary Geometrical Optics 77 The Limits of Resolution 79 Different Types of Microscopy 82 5.4.1 Bright field microscopy 82 Contents xi 5.4.2 5.4.3 5.4.4 5.4.5 Dark field microscopy 82 Phase contrast microscopy 82 Fluorescence microscopy 83 Polarising microscopy 84 6. Electron Microscopy 86 6.1 6.2 6.3 6.4 6.5 6.6 6.7 6.8 6.9 Introduction 86 Electron Optics 86 The Transmission Electron Microscope (TEM) 87 The Scanning Electron Microscope (SEM) 88 Preparation of the Specimen for Electron Microscopy 90 Image Reconstruction 91 Electron Diffraction 92 The Tunnelling Electron Microscope 92 Atomic Force Microscope 93 7. X-ray Crystallography 7.1 7.2 7.3 7.4 7.5 7.6 7.7 7.8 Introduction 95 Crystals and Symmetries 95 Crystal Systems 96 Point Groups and Space Groups 97 Growth of Crystals of Biological Molecules 99 X-ray Diffraction 103 X-ray Data Collection 105 Structure Solution 106 7.8.1 7.8.2 7.8.3 7.8.4 The heavy atom or multiple isomorphous replacement (MIR) method 107 Direct methods 108 Molecular replacement 108 The anomalous scattering technique 108 7.9 Refinement of the Structure 108 7.9.1 7.9.2 7.9.3 Least squares methods 108 Constrained-restrained refinement 109 Computer graphics in refinement 110 7.10 Note on the Resolution of an X-ray Structure 111 8. 95 NMR Spectroscopy 112 8.1 8.2 8.3 8.4 8.5 Introduction 112 Basic Principles of NMR 112 NMR Theory and Experiment 114 Classical Description of NMR 115 NMR Parameters 117 8.5.1 8.5.2 8.5.3 8.5.4 8.5.5 Chemical shift 117 Intensity 118 Line width 119 Relaxation parameters 119 Spin-spin coupling 120 8.6 The Nuclear Overhauser Effect 122 xii Contents 8.7 NMR Applications in Chemistry 122 8.7.1 8.7.2 8.7.3 Chemical shift 122 Spin-spin coupling 123 NMR 124 8.8 NMR Applications in Biochemistry and Biophysics 124 8.8.1 8.8.2 8.8.3 8.8.4 8.8.5 Concentration measurement 125 pH titration 125 Conformation of biomolecules 125 Two-Dimensional NMR 126 Determination of macromolecular structure 128 8.9. NMR in Medicine 128 9. Molecular Modelling 131 9.1 9.2 Introduction 1 3 1 Generating the Model 1 3 1 9.2.1 9.2.2 9.2.3 9.2.4 Building the structure of 1 3 2 Building protein structure 1 3 3 Building nucleic acid structure 1 3 6 Displaying and altering the generated model 1 3 7 9.3 Optimising the Model 1 3 8 10. Macromolecular Structure 144 10.1 10.2 Introduction 144 Nucleic Acid Structure 144 10.2.1 10.2.2 10.2.3 10.2.4 10.2.5 10.2.6 The chemical structure of nucleic acids 145 Conformational possibilities of monomers and polymers 147 The double helical structure of DNA 148 Polymorphism of DNA 151 DNA supercoiling and unusual DNA structures 154 The structure of transfer RNA 158 10.3 Protein Structure 161 10.3.1 10.3.2 10.3.3 10.3.4 10.3.5 Amino acids and the primary structure of proteins 161 The peptide bond and secondary structure of proteins 161 Tertiary structure—supersecondary and domain structure 169 Quaternary structure 170 Virus structure 172 11. Energy Pathways in Biology 174 11.1 11.2 11.3 11.4 11.5 11.6 Introduction 174 Free Energy 174 Coupled Reactions 176 Group Transfer Potential 177 Role of Pyridine Nucleotides 178 Photosynthesis 179 11.6.1 11.6.2 11.6.3 Phostosystem I 183 Photosystem II 185 Photophosphorylation and carbon fixation 186 11.7 Energy Conversion Pathways 187 Contents xiii 11.7.1 11.7.2 11.7.3 11.7.4 Oxidation 187 Glycolysis 187 The Krebs cycle 188 The respiratory chain 190 11.8 Membrane Transport 194 11.8.1 11.8.2 Active transport 195 Chemi-osmotic theory—passive transport 196 12. Biomechanics 199 12.1 12.2 12.2.1 Introduction 199 StriatedMuscles 199 Contractile proteins 201 12.3 Mechanical Properties of Muscles 202 12.3.1 12.3.2 Contraction mechanism 203 Role of ions 204 12.4 Biomechanics of the Cardiovascular System 205 12.4.1 12.4.2 12.4.3 Bloodpressure 205 Electrical activity during the heartbeat 207 Electrocardiography 208 13. Neurobiophysics 210 13.1 13.2 Introduction 210 The Nervous System 210 13.2.1 Synapse 212 13.3 Physics of Membrane Potentials 213 13.3.1 13.3.2 Membrane potential due to diffusion 213 Voltage Clamp 215 13.4 Sensory Mechanisms—The Eye 217 13.4.1 13.4.2 13.4.3 13.4.4 13.4.5 The visual receptor 218 Electrical activity andvisual generator potentials 223 Optical defects of the eye 224 Neural aspects of vision 224 Visual communications, bioluminescence 226 13.5 Physical Aspects of Hearing 227 13.5.1 13.5.2 13.5.3 The Ear 227 Elementary acoustics 228 Theories of hearing 230 13.6 Signal Transduction 230 13.6.1 13.6.2 Mode of transport 231 Signal transduction in the cell 231 14. Origin and Evolution of Life 232 14.1 14.2 14.3 14.4 Introduction 232 Prebiotic Earth 233 Theories of Origin andEvolution of Life 234 Laboratory Experiments on the Formation of Small Molecules 235 14.4.1 14.4.2 Prebiotic synthesis of amino acids—Miller andUrey’s experiment 235 Prebiotic synthesis of purines, pyrimidines and nucleosides 237 xiv Contents 14.4.3 14.4.4 14.4.5 14.4.6 Sugars 2 3 7 Nucleotide synthesis 2 3 8 Optical activity 2 3 9 Polymerisation 240 FURTHER READING 243 INDEX 247