Johnson / Holt / Ackers Biothermodynamics, Part B

1. Auflage 2009
ISBN: 978-0-08-088781-4
Verlag: Elsevier Science & Techn.
Format: EPUB
Kopierschutz: 6 - ePub Watermark

E-Book, Englisch, 676 Seiten

The use of thermodynamics in biological research can be equated to an 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. This volume presents sophisticated methods for estimating the thermodynamic parameters of specific protein-protein, protein-DNA and small molecule interactions.
* Elucidates the relationships between structure and energetics and their applications to molecular design, aiding researchers in the design of medically important molecules * Provides a 'must-have' methods volume that keeps MIE buyers and online subscribers up-to-date with the latest research * Offers step-by-step lab instructions, including necessary equipment, from a global research community

Autoren/Hrsg.

Johnson, Michael L.

Holt, Jo M.

Ackers, Gary K.

Inhaltsverzeichnis

1;Front Cover;1
2;Methods in Enzymology;4
3;Copyright Page;5
4;Contents;6
5;Contributors;14
6;Preface;20
8;Chapter 1: Using NMR-Detected Backbone Amide 1H Exchange to assess Macromolecular Crowding Effects on Globular-Protein Stability;50
8.1;1. Introduction;51
8.2;2. Globular Protein Stability;52
8.3;3. Mechanism and Limits of Amide 1H Exchange;52
8.4;4. Requirements for Candidate Systems;55
8.5;5. Preliminary Experiments;57
8.6;6. A Protocol for Amide 1H Exchange;63
8.7;7. Summary and Future Directions;64
8.8;Acknowledgments;65
8.9;References;65
9;Chapter 2: Fluorescence Spectroscopy in Thermodynamic and Kinetic Analysis of pH-Dependent Membrane Protein Insertion;68
9.1;1. Introduction: Co-Translational Versus Post-Translational Membrane Protein Insertion;70
9.2;2. Challenges of Thermodynamic Analysis of Membrane Protein Folding/Insertion;71
9.3;3. FCS and Protein-Membrane Interactions;72
9.4;4. Thermodynamic Schemes for Analysis of Membrane Partitioning;77
9.5;5. Kinetic Analysis of Membrane Protein Insertion;81
9.6;6 Perspectives: Annexin B12 as a Model for Thermodynamic and Kinetic Analysis of Membrane Protein Insertion, Folding and Misfolding;87
9.7;Acknowledgments;89
9.8;References;89
10;Chapter 3: Evaluating the Energy-Dependent ``Binding´´ in the Early Stage of Protein Import into Chloroplasts;92
10.1;1. Introduction;93
10.2;2. The In Vitro Chloroplastic Import Assay Using Recombinant Precursor Proteins;94
10.3;3. Limited Proteolysis of Docked Precursor Proteins;102
10.4;4. The Behavior of Transit Peptide During the Transition;108
10.5;5. Conclusions;111
10.6;Acknowledgments;111
10.7;References;111
11;Chapter 4: Use of DNA Length Variation to Detect Periodicities in Positively Cooperative, Nonspecific Binding;114
11.1;1. Introduction;115
11.2;2. Protein and DNA Preparations;117
11.3;3. Stoichiometry Analyses;117
11.4;4. Affinity and Cooperativity as Functions of DNA Length;124
11.5;Acknowledgments;127
11.6;References;127
12;Chapter 5: The Impact of Ions on Allosteric Functions in Human Liver Pyruvate Kinase;132
12.1;1. Introduction;133
12.2;2. General Strategy to Assess Allosteric Coupling;135
12.3;3. PYK Assay;137
12.4;4. Buffers;140
12.5;5. Divalent Cation;142
12.6;6. Monovalent Cation;144
12.7;7. Anion;149
12.8;8. Concluding Remarks;152
12.9;Acknowledgments;154
12.10;References;154
13;Chapter 6: Conformational Stability of Cytochrome c Probed by Optical Spectroscopy;158
13.1;1. Introduction;159
13.2;2. Basic Theory of Absorption and Circular Dichroism Spectroscopy;162
13.3;3. Secondary Structure Analysis of Cytochrome c Using Ultra-Violet Circular Dichroism Spectroscopy;166
13.4;4. Visible CD and Absorption Spectroscopy of Native Cytochrome c;170
13.5;5. Nonnative States of Ferricytochrome c Probed by Visible CD and Absorption Spectroscopy;180
13.6;6. Summary and Outlook;197
13.7;References;198
14;Chapter 7: Examining Ion Channel Properties Using Free-Energy Methods;204
14.1;1. Introduction;205
14.2;2. Free-Energy Calculations ;206
14.3;3. Thermodynamic Integration;208
14.4;4. Free-Energy Perturbation;209
14.5;5. Umbrella Sampling;211
14.6;6. Adaptive Biasing Force;213
14.7;7. Metadynamics;216
14.8;8. Applications of Free-Energy Methods to Study Ion Channel Properties;218
14.9;9. Conclusions and Future Outlook;223
14.10;Acknowledgments;224
14.11;References;224
15;Chapter 8: Examining Cooperative Gating Phenomena in Voltage-Dependent Potassium Channels: Taking the Energetic Approach;228
15.1;1. Introduction;229
15.2;2. High-Order Thermodynamic Mutant Cycle Coupling Analysis;230
15.3;3. The Voltage-Activated Potassium Channel Allosteric Model System;237
15.4;4. Deriving a Hill Coefficient for Assessing Cooperativity in Voltage-Dependent Ion Channels ;242
15.5;5. Direct Analysis of Cooperativity in Multisubunit Allosteric Proteins;245
15.6;6. Long-Range Energetic Coupling Mediated Through Allosteric Communication Trajectories;251
15.7;7. Concluding Remarks;256
15.8;Acknowledgments;256
15.9;References;256
16;Chapter 9: Thermal Stability of Collagen Triple Helix;260
16.1;1. Introduction;261
16.2;2. Methods;263
16.3;References;280
17;Chapter 10: Electrostatic Contributions to the Stabilities of Native Proteins and Amyloid Complexes;282
17.1;1. Introduction;283
17.2;2. Practical Aspects of pKa Measurements by NMR;285
17.3;3. Interpreting pKa Values in Terms of Stability;289
17.4;4. Importance of the Reference (Unfolded) State;289
17.5;5. Results from Globular Proteins;289
17.6;6. Results from Coiled Coils;290
17.7;7. Comparison of NMR and Crystallographic Results;291
17.8;8. Comparison of NMR and Mutagenesis: Nonadditivity of Ion Pairs;292
17.9;9. Improving Structure-Based Modeling of pKa Values;293
17.10;10. Results with Micelle-Bound Proteins;294
17.11;11. Results from Fibrillization Kinetics;298
17.12;12. Conclusion;302
17.13;Acknowledgments;303
17.14;References;303
18;Chapter 11: Kinetics of Allosteric Activation;308
18.1;1. Linkage;308
18.2;2. Allosteric Activation at Steady State;310
18.3;3. Different Types of Activation (Type Ia, Type Ib, and Type II);315
18.4;4. Concluding Remarks;318
18.5;Acknowledgment;319
18.6;References;319
19;Chapter 12: Thermodynamics of the Protein Translocation;322
19.1;1. Introduction;323
19.2;2. Example 1: SecA Nucleotide Binding;327
19.3;3. Example 2: Probing SecB:Substrate Interactions;332
19.4;4. Concluding Remarks;337
19.5;References;338
20;Chapter 13: Thermodynamic Analysis of the Structure-Function Relationship in the Total DNA-Binding Site of Enzyme-DNA Complexes;342
20.1;1. Introduction;343
20.2;2. Thermodynamic Bases of Quantitative Equilibrium Spectroscopic Titrations;345
20.3;3. Anatomy of the Total DNA-Binding Site in the PriA Helicase-ssDNA Complex;351
20.4;4. Structure-Function Relationship in the Total ssDNA-Binding Site of the DNA Repair Pol X From ASFV;366
20.5;Acknowledgments;371
20.6;References;371
21;Chapter 14: Equilibrium and Kinetic Approaches for Studying Oligomeric Protein Folding;374
21.1;1. Introduction;375
21.2;2. Methods to Monitor Folding and Association;376
21.3;3. Equilibrium Studies;385
21.4;4. Kinetic Studies;392
21.5;Acknowledgments;403
21.6;References;403
22;Chapter 15: Methods for Quantifying T cell Receptor Binding Affinities and Thermodynamics;408
22.1;1. Introduction;409
22.2;2. Isothermal Titration Calorimetry of TCR-Peptide/MHC Interactions;411
22.3;3. Surface Plasmon Resonance Studies of TCR-Peptide/MHC Interactions;416
22.4;4. Fluorescence Anisotropy as a Tool for Characterizing TCR-Peptide/MHC Interactions;422
22.5;5. Concluding Remarks;427
22.6;Acknowledgments;427
22.7;References;427
23;Chapter 16: Thermodynamic and Kinetic Analysis of Bromodomain-Histone Interactions;432
23.1;1. Introduction;433
23.2;2. Fluorescence Anisotropy Theory and Concepts;433
23.3;3. Developing Binding Models for the Analysis of Fluorescence Anisotropy Data;435
23.4;4. Experimental Considerations in Designing Fluorescence Anisotropy Assays;439
23.5;5. Preparation of Histone and Bromodomain Samples;440
23.6;6. Fluorescence Anisotropy Measurements;441
23.7;7. Kinetic Analysis;444
23.8;8. Determination of Thermodynamic Parameters;448
23.9;9. Thermodynamic Measurements;449
23.10;10. Developing a Binding Model;452
23.11;11. Concluding Remarks;454
23.12;Acknowledgments;454
23.13;References;454
24;Chapter 17: Thermodynamics of 2-Cys Peroxiredoxin Assembly Determined by Isothermal Titration Calorimetry;458
24.1;1. Introduction;459
24.2;2. Dimer-Decamer Equilibrium;461
24.3;3. Isothermal Titration Calorimetry-General Concepts;464
24.4;4. ITC Dilution Experiments;465
24.5;5. Material and Instruments;467
24.6;6. Experimental Procedure;467
24.7;7. Results, Data Analysis, and Discussion;472
24.8;8. Conclusions;477
24.9;Acknowledgments;477
24.10;References;477
25;Chapter 18: Protein-Lipid Interactions: Role of Membrane Plasticity and Lipid Specificity on Peripheral Protein Interactions;480
25.1;1. Introduction;481
25.2;2. Defining Protein-Lipid Interactions;482
25.3;3. Selective Partitioning and Lipid Activities;483
25.4;4. Protein-Protein Interactions at the Membrane Surface;484
25.5;5. Measuring Protein-Lipid Interactions;486
25.6;6. Modeling of Protein-Lipid Interactions;488
25.7;7. Synopsis;497
25.8;Acknowledgments;499
25.9;References;500
26;Chapter 19: Predicting pKa Values with Continuous Constant pH Molecular Dynamics;504
26.1;1. Introduction;505
26.2;2. Theoretical Methods for pKa Predictions;506
26.3;3. Predicting Protein pKas with REX-CPHMD Simulations;514
26.4;4. Conclusions;519
26.5;Acknowledgment;520
26.6;References;520
27;Chapter 20: Unfolding Thermodynamics of DNA Intramolecular Complexes Involving Joined Triple- and Double-Helical Motifs;526
27.1;1. Introduction;527
27.2;2. Materials and Methods;530
27.3;3. Results and Discussion;534
27.4;4. Conclusions;548
27.5;Acknowledgments;548
27.6;References;549
28;Chapter 21: Thermodynamics and Conformational Change Governing Domain-Domain Interactions of Calmodulin;552
28.1;1. Introduction;553
28.2;2. Overexpression and Purification of rCaM Fragments;556
28.3;3. Calcium-Binding Properties of N-Domain CaM Fragments;556
28.4;4. Tertiary Constraints of N-Domain CaM Fragments;561
28.5;5. Tertiary Conformation of N-Domain CaM Fragments;565
28.6;6. High-Resolution Studies of N-Domain CaM Fragments;568
28.7;7. Conclusions;571
28.8;Acknowledgments;573
28.9;References;574
29;Chapter 22: Use of Pressure Perturbation Calorimetry to Characterize the Volumetric Properties of Proteins;576
29.1;1. Introduction;577
29.2;2. Determination of the Coefficient of Thermal Expansion (aPr) Using PPC;580
29.3;3. Sample Preparation;582
29.4;4. Derivation of a Two-State Model for Analysis of PPC Data;584
29.5;5. Practical Considerations;588
29.6;6. Implications of Two-State Model for Future PPC Experiments ;594
29.7;References;594
30;Chapter 23: Solvent Denaturation of Proteins and Interpretations of the m Value;598
30.1;1. Introduction;598
30.2;2. Protein Unfolding or Denaturation ;599
30.3;3. Linear Extrapolation Method;604
30.4;4. DeltaG(H2O): Conformational Stability;605
30.5;5. The m Value;607
30.6;6. Concluding Remarks;611
30.7;Acknowledgments;612
30.8;References;612
31;Chapter 24: Measuring Cotranslational Folding of Nascent Polypeptide Chains on Ribosomes;616
31.1;1. Introduction;617
31.2;2. Translation and the Ribosome:Nascent Chain (RNC) Complex;619
31.3;3. General Approaches for Generating Stalled RNC Complexes;621
31.4;4. Methods for Preparing RNC Complexes;626
31.5;5. Biophysical Studies with RNC Complexes;628
31.6;6. Measuring Nascent Chain Cotranslational Folding and Rigidity by Limited Protease Digestion;632
31.7;7. Future Directions;633
31.8;References;634
32;Author Index;640
33;Subject Index;652
34;Color Plates;664