Index -I-
Contents
PREFACE V CHAPTER I 1 Introduction 1 1.1 Binding of drugs to Polynucleotides 41.1.1 Binding process I: Intercalation 4
1.1.2 Binding process II: External binding 7
1.1.3 Electrostatic Binding 9
1.1.4 Groove Binding 10
1.2 Studies on DNA/drugs interactions 12
1.2.1 The x-ray technique and theoretical calculations 12
1.2.2 Solution techniques 13
1.2.3 Binding titrations and site size determination 15
1.2.4 Kinetic analysis of drug/polynucleotide binding 17
CHAPTER II 19
Materials and Methods 19
2.1 Materials 19
2.1.1 Dyes 19
2.1.2 Nucleic acids 20
2.1.3 Reaction media 21
2.2 Methods 22
2.2.1 Spectrophotometry and spectrofluorometry 22
2.2.2 Circular Dichroism 22
2.2.3 Viscosity 22
2.2.4 Kinetics 22
CHAPTER III 27
Alkaloids: Coralyne interaction with DNA and RNA 27 3.1 Dye self-aggregation 29
3.1.2 Spectral analysis of aggregation 29
3.1.3 Kinetic analysis of self-aggregation 32
3.2 Coralyne/nucleic acids interaction: Equilibria 34
3.2.1 Absorbance titrations 34
3.2.2 Fluorescence titrations 36
3.2.3 Salt concentration dependence of equilibria 41
3.2.4 Solvent effect on the nucleic acids/coralyne equilibria 42
3.3 CD measurements 43
3.4 Viscometric measurements 45 3.5 DNA/coralyne interaction: Kinetics 46
3.6 Discussion 48
CHAPTER IV 53
Acridines: D and ZnD interaction with DNA 53 4.1 Spectral analysis of aggregation 54
4.2 NMR measurements 55
Index
-II
4.3.1 Equilibria 56
4.3.2 Salt concentration dependence of equilibria 59
4.3.3 Dye displacement Assay 60
4.3.4 Circular dichroism measurements 61
4.4 Supercoiled DNA unwinding 62 4.5 Kinetics of DNA/D and DNA/ZnD systems 63
4.6 Discussion 65
CHAPTER V 69
Cyanines: BO and TO interaction with DNA 69
5.1 Dye self-aggregation 70
5.1.1 Lambert & Beer’s law 70
5.1.2 Spectral analysis of aggregation 70
5.1.3 Kinetic analysis of TO self-aggregation 73
5.2 Equilibria 75
5.2.1 The DNA/BO system 75
5.2.2 The DNA/TO system 80
5.2.3 Salt concentration dependence of equilibria 83
5.2.4 Temperature dependence of equilibria 83
5.2.5 Interaction with synthetic polynucleotides 85
5.3 Kinetics 85
3.3.1 The DNA/BO system 85
3.3.2 The DNA/TO system 88
3.3.3 Amplitude analysis 89
5.4 Discussion 90
CHAPTER VI 97
Cyanines: T307 interaction with DNA 97
6.1 Dye self-aggregation 98
6.1.1 Thermodynamics of self-aggregation 98
6.1.2 Kinetics of self-aggregation 99
6.1.3 Effect of SDS micelles 101
6.1.4 Light scattering experiments 102
6.2 Dye interaction with DNA 103
6.3 CD measurements 106
6.4 Kinetics 107
APPENDIX I
Derivation of equations for the analysis of dye self-aggregation
111 APPENDIX II
Determination of the equilibrium constant for complex formation when the ligand is a polynucleotide
115
APPENDIX III
Determination of equilibrium constant for complex formation and of site size using the Scatchard and McGhee and von Hippel equations
119
APPENDIX IV
Record equation for the analysis of equilibrium constant dependence on salt concentration
Index
-III-APPENDIX V
Determination of site size and equilibrium constant for complex formation according to the Schwarz theory
125
APPENDIX VI
Derivation of relaxation times equation for a three step series mechanism
129
APPENDIX VII
Amplitude analysis of kinetic relaxation curves 133
References 137 Publications and congresses 153
Index
