The aim of this book is to give a comprehensive description of the basic methods used in the ultraviolet spectroscopy of proteins, to discuss new trends and development of these methods, and to analyze their different applications in the study of various aspects of protein structure and dynamics. Ultraviolet spectroscopy is one of the oldest and most popular methods in the field of biochemistry and molecular biophysics. At present, it is difficult to imagine the biochemical laboratory without a recording spectrophotometer or spectrofluorimeter. There are several hundreds of publications directly devoted to protein ultraviolet spectroscopy and in a great number of studies UV spectroscopic methods are used for the structural analysis of different proteins. Meanwhile a unified description of the theoretical basis of the methods, experimental techniques, data analysis, and generalization of results obtained in solving the specific problems of protein structure are lacking. There are three reasons for which a monograph on ultraviolet spectroscopy is needed today. Firstly, there has been significant growth in facilities of experimental technique, its precision, and versatility associated with computer data analysts. This new technique is available to a wide circle of scientists engaged in the field of protein research. Most of them are not spectroscopists and, thus, there is a need for a conceivable and precise source of information on how to use this method and what kind of data it should provide.

Inhaltsangabe1. Spectroscopic Properties of Protein Chromophores.- 1.1 Basic Principles and Definitions of Light Absorption and Emission Spectroscopy.- 1.1.1 Absorption.- 1.1.2 Emission.- 1.2 Light Absorption by the Amide Chromophore.- 1.3 The Absorption Spectra of Amino Acid Residues and Their Analogs.- 1.3.1 Phenylalanine.- 1.3.2 Tyrosine.- 1.3.3 Tryptophan.- 1.3.4 Other Protein Chromophores.- 1.4 Emission Properties of Aromatic Amino Acids.- 1.4.1 Phenylalanine.- 1.4.2 Tyrosine.- 1.4.3 Tryptophan.- 1.5 Conclusions.- 2. Display of Intramolecular and Intermolecular Interactions in Electronic Spectra of Amino Acids and Proteins.- 2.1 Spectroscopic Analysis of the Environmental Polarity and Polarizability Effects.- 2.2 Spectroscopic Manifestation of the Hydrogen Bond.- 2.3 Substitution and Charge Effects.- 2.4 Charge-Transfer Complexes.- 2.5 Broadening of Electronic Spectra.- 2.6 Excited State Processes.- 2.6.1 Solvent Relaxation.- 2.6.2 Exciplexes.- 2.6.3 Excited State Proton Transfer.- 2.6.4 Excited State Electron Transfer.- 2.6.5 On the Nature of the Emitting State of Indole and Tryptophan.- 2.7 Conclusions.- 3. Difference Spectra of Proteins -.- 3.1 Informational Significance of Difference Spectra.- 3.2 Studies of Protein Denaturation.- 3.3 Functional Transformations and Association of Proteins.- 3.3.1 Conformational Changes Under Activation of Precursor Proteins.- 3.3.2 Formation of Enzyme-Substrate and Enzyme-Inhibitor Complexes. Studies on Mechanisms of Catalytic Activity.- 3.3.3 Interaction of Proteins with Small Molecules and Ions.- 3.3.4 Association of Subunits and Formation of Supermolecular Structures.- 3.4 Solvent Perturbation Difference Spectra and Studies in Surface Topography.- of Protein Molecules.- 3.5 Protein-Model Difference Spectra.- 3.6 Conclusions.- 4. Thermal Perturbation Difference Spectroscopy and Temperature-Dependent Conformational Transitions of Proteins.- 4.1 Characteristics of Tyrosine, Tryptophan, and Phenylalanine Spectra and Their Origin.- 4.1.1 Quantitative Analysis of the Experimental Data. Account of Correction Factors.- 4.1.2 TPDS of Tryptophan, Tyrosine, and Phenylalanine.- 4.1.3 Origin of Thermal Perturbation Difference Spectra.- 4.1.4 On the Nature of Longwave Shift with the Temperature Rise.- 4.2 Studies of Thermal Perturbation of Tyrosine, Tryptophan, and Phenylalanine Residues in Proteins.- 4.2.1 Drop in Intensity of TPDS.- 4.2.2 Longwave Shift of TPDS Maxima.- 4.2.3 Absence of Tyrosine Maxima at 287-289 nm.- 4.2.4 Presence of TPDS Maxima in the Region of 300-307 nm.- 4.3 TPDS and Protein Conformational Transitions Depending on Temperature.- and pH of the Medium.- 4.4 Conclusions.- 5. Derivative Spectroscopy of Aromatic Amino Acids and Proteins.- 5.1 The Theoretical Grounds.- 5.2 Derivative Spectra of Tryptophan, Tyrosine, and Phenylalanine.- 5.3 Influence of Solvents on the Derivative Spectra of Aromatic Amino Acids.- 5.4 Analysis of Chromophore Environment in Proteins.- 5.4.1 The State of Phenvlalanine Residues.- 5.4.2 The State of Tyrosine and Tryptophan.- 5.5 Studies of Conformational Transitions in Proteins. Difference-Derivative Spectroscopy.- 5.6 Studies on Broadening of Absorption Spectra.- 5.7 Conclusions.- 6. Spectrophotometric Titration of Proteins.- 6.1 The Spectrophotometric Titration Method.- 6.2 Titration of Tyrosine Residues at Alkaline pH.- 6.3 Conclusions.- 7. Fluorescence Molecular Relaxation Spectroscopy.- 7.1 Relaxational Shift of the Fluorescence Spectra.- 7.2 Time-Resolved Spectroscopy.- 7.3 Edge Excitation Fluorescence Spectroscopy.- 7.3.1 Physical Principles.- 7.3.2 The Effect of Selective Excitation on Fluorescence Spectra of Indole and Tryptophan.- 7.3.3 The Structural Relaxation of Indolic Chromophore in Proteins.- 7.4 Conclusions.- 8. Fluorescence Quenching.- 8.1 Effects of External Difiusional Quenchers.- 8.2 Quenching by Protein Internal Groups and Its Temperature Dependence.- 8.3 Conclusions.- 9. Nonradiative Transfer of Electronic Excitation Energy.- 9.1 General