In the past several years, there has been an explosion in the ability of biologists, molecular biologists and biochemists to collect vast amounts of data on their systems. This volume presents sophisticated methods for estimating the thermodynamic parameters of specific protein-protein, protein-DNA and small molecule interactions. The use of thermodynamics in biological research is used as an "e;energy book-keeping system.? While the structure and function of a molecule is important, it is equally important to know what drives the energy force. These methods look to answer: What are the sources of energy that drive the function? Which of the pathways are of biological significance? As the base of macromolecular structures continues to expand through powerful techniques of molecular biology, such as X-ray crystal data and spectroscopy methods, the importance of tested and reliable methods for answering these questions will continue to expand as well.
Front Cover 1
Methods in Enzymology Biothermodynamics, Part A 4
Copyright Page 5
Contents 6
Contributors 12
Preface 16
Volumes in Series 18
Chapter 1: Practical Approaches to Protein Folding and Assembly: Spectroscopic Strategies in Thermodynamics and Kinetics 46
1. Introduction 47
2. Equilibrium Unfolding 48
3. Measuring Folding Kinetics 66
References 81
Chapter 2: Using Thermodynamics to Understand Progesterone Receptor Function: Method and Theory 86
1. Introduction 87
2. Assessing Protein Functional and Structural Homogeneity 88
3. Dissecting Linked Assembly Reactions 91
4. Analysis and Dissection of Natural Promoters 99
5. Measuring the Energetics of Coactivator Recruitment 107
6. Correlation to Biological Function 109
7. Conclusions and Future Directions 112
Acknowledgments 113
References 113
Chapter 3: Direct Quantitation of Mg2+-RNA Interactions by Use of a Fluorescent Dye 116
1. Introduction 117
2. General Principles 118
3. Ion-Binding Properties of HQS 123
4. Preparation of Solutions and Reagents 126
5. Instrumentation and Data Collection Protocols 129
6. Data Analysis 133
7. Controls and Further Considerations 135
Acknowledgments 137
References 137
Chapter 4: Analysis of Repeat-Protein Folding Using Nearest-Neighbor Statistical Mechanical Models 140
1. Historical Overview of Ising Models and Motivation for the Present Review 141
2. Linear Repeat Proteins and Their Connection to Linear Ising Models 142
3. Formulating a Homopolymer Partition Function and the Zipper Approximation 145
4. Matrix Approach: Homopolymers 149
5. Matrix Approach: Heteropolymers 154
6. Solvability Criteria for Ising Models Applied to Repeat-Protein Folding 156
7. Matrix Homopolymer Analysis of Consensus TPR Folding 160
8. Matrix Heteropolymer Analysis of Consensus Ankyrin Repeat Folding 164
9. Summary and Future Directions 168
Acknowledgments 169
References 169
Chapter 5: Isothermal Titration Calorimetry: General Formalism using Binding Polynomials 172
1. Introduction 173
2. The Binding Polynomial 174
3. Microscopic Constants and Cooperativity 176
4. Independent or Cooperative Binding? 177
5. Analysis of ITC Data using Binding Polynomials 178
6. A Typical Case: Macromolecule with Two Ligand-Binding Sites 180
7. Data Analysis 182
8. Data Interpretation 186
9. An Experimental Example 191
10. Experimental Situations from the Literature 192
11. Macromolecule with Three Ligand-Binding Sites 195
12. Conclusions 195
Appendix 196
Acknowledgments 199
References 199
Chapter 6: Kinetic and Equilibrium Analysis of the Myosin ATPase 202
1. Introduction 203
2. Reagents and Equipment used for all Assays 204
3. Steady-State ATPase Activity of Myosin 206
4. Steady-State Measurement of Actomyosin Binding Affinities 211
5. Transient Kinetic Analysis of the Individual ATPase Cycle Transitions 215
6. Kinetic Simulations 233
Acknowledgments 234
References 235
Chapter 7: The Hill Coefficient: Inadequate Resolution of Cooperativity in Human Hemoglobin 238
1. Introduction 239
2. Cooperativity and Intrinsic Binding 239
3. The Macroscopic Binding Isotherm 242
4. The Hill Coefficient 245
5. Microscopic Cooperativity in Hemoglobin 250
6. Summary 256
References 257
Chapter 8: Methods for Measuring the Thermodynamic Stability of Membrane Proteins 258
1. Introduction 259
2. Two Classes of Membrane Proteins 260
3. Methods for Measuring Transmembrane Domain Oligomer Stability 261
4. Methods for Measuring Multipass alpha-Helical Membrane Protein Stability 264
5. Methods to Study the Stability of beta-barrel Membrane Proteins 267
6. A Few Salient Results on Forces that Stabilize Membrane Proteins 272
7. Conclusion and Outlook 276
Acknowledgments 277
References 277
Chapter 9: NMR Analysis of Dynein Light Chain Dimerization and Interactions with Diverse Ligands 282
1. NMR Methodology 283
2. Monomer-dimer Equilibrium Coupled to Electrostatics 286
3. Dimerization is Coupled to Ligand Binding 291
4. Folding is Coupled to Binding 292
5. Allostery in LC8 296
6. Summary 300
References 301
Chapter 10: Characterization of Parvalbumin and Polcalcin Divalent Ion Binding by Isothermal Titration Calorimetry 304
1. Introduction 305
2. Practical Aspects of Data Collection 307
3. Illustrative Global ITC Analyses of Divalent Ion Binding 326
4. Conclusion 340
Acknowledgments 340
References 340
Chapter 11: Energetic Profiling of Protein Folds 344
1. Introduction 345
2. Modeling the Native State Ensemble of Proteins using Statistical Thermodynamics 346
3. Energetic Profiles of Proteins Derived from Thermodynamics of the Native State Ensemble 349
4. Principal Components Analysis of Energetic Profile Space 351
5. Energetic Profiles are Conserved between Homologous Proteins 353
6. Direct Alignment of Energetic Profiles Based on a Variant of the CE Algorithm 360
7. CE Algorithm Described for Structure Coordinates 361
8. Necessary Deviations from the CE Algorithm to Accommodate Energetic Profiles 362
9. Towards a Thermodynamic Homology of Fold Space: Clustering Energetic Profiles using STEPH 363
10. Energetic Profiles Provide a Vehicle to Discover Conserved Substructures in the Absence of Known Homology 366
11. Conclusion 368
Acknowledgments 370
References 370
Chapter 12: Model Membrane Thermodynamics and Lateral Distribution of Cholesterol: From Experimental Data to Monte Carlo Simulation 374
1. Introduction 375
2. Materials and Methods 376
3. Result and Discussion 383
4. Concluding Remarks 407
Acknowledgments 407
References 408
Chapter 13: Thinking Inside the Box: Designing, Implementing, and Interpreting Thermodynamic Cycles to Dissect Cooperativity in RNA and DNA Folding 410
1. Introduction 411
2. Folding Cooperativity Defined 412
3. Thermodynamic Boxes: Design, Implementation, and Interpretation 414
4. Thermodynamic Cubes: Design, Implementation, and Interpretation 419
5. Examples of Cooperativity in RNA 421
6. Measuring Thermodynamic Parameters by UV Melting 424
7. Concluding Remarks 435
Acknowledgments 436
References 436
Chapter 14: The Thermodynamics of Virus Capsid Assembly 440
1. Introduction 441
2. The Structural Basis of Capsid Stability 442
3. Analysis of Capsid Stability 446
4. Applications of Thermodynamic Evaluation of Virus Capsid Stability 453
5. Concluding Remarks 459
References 459
Chapter 15: Extracting Equilibrium Constants from Kinetically Limited Reacting Systems 464
1. Introduction 465
2. Methods 466
3. Simulation and Analysis of Dimerization 466
4. Kinetically Mediated Dimerization 473
5. A Stepwise Approach 481
6. Final Thoughts 487
Acknowledgments 488
References 488
Author Index 492
Subject Index 506
Color Plate Section 512
Erscheint lt. Verlag | 14.3.2009 |
---|---|
Sprache | englisch |
Themenwelt | Naturwissenschaften ► Biologie ► Biochemie |
Naturwissenschaften ► Biologie ► Genetik / Molekularbiologie | |
Naturwissenschaften ► Chemie | |
Naturwissenschaften ► Physik / Astronomie ► Angewandte Physik | |
Naturwissenschaften ► Physik / Astronomie ► Thermodynamik | |
Technik | |
ISBN-10 | 0-08-092340-2 / 0080923402 |
ISBN-13 | 978-0-08-092340-6 / 9780080923406 |
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