Cadenas / Packer Thiol Redox Transitions in Cell Signaling, Part A

Chemistry and Biochemistry of Low Molecular Weight and Protein Thiols
1. Auflage 2010
ISBN: 978-0-12-381346-6
Verlag: Elsevier Science & Techn.
Format: EPUB
Kopierschutz: 6 - ePub Watermark

E-Book, Englisch, Band 473, 416 Seiten

Reihe: Methods in Enzymology

This volume, along with its companion (volume 475), presents methods and protocols dealing with thiol oxidation-reduction reactions and their implications as they relate to cell signaling. This first installment of Cadenas and Packer's two-volume treatment specifically deals with glutathionylation and dethiolation, and peroxide removal by peroxiredoxins/thioredoxins and glutathione peroxidases. The critically acclaimed laboratory standard for 40 years, Methods in Enzymology is one of the most highly respected publications in the field of biochemistry. Since 1955, each volume has been eagerly awaited, frequently consulted, and praised by researchers and reviewers alike. Over 450 volumes have been published to date, and much of the material is relevant even today--truly an essential publication for researchers in all fields of life sciences.
*Along with companion volume, provides a full overview of techniques necessary to the study of thiol redox in relation to cell signaling
* Gathers tried and tested techniques from global labs, offering both new and tried-and-true methods
* Relevant background and reference information given for procedures can be used as a guide to developing protocols in a number of disciplines

Autoren/Hrsg.

Cadenas, Enrique

Packer, Lester

Inhaltsverzeichnis

1;Front Cover;1
2;Methods in Enzymology: Thiol Redox Transitions in Cell Signaling, Part A;4
3;Copyright Page;5
4;Contents;6
5;Contributors;12
6;Preface;20
7;Volumes in Series;22
8;Chapter 1: Changing Paradigms in Thiology: From Antioxidant Defense Toward Redox Regulation ;50
8.1;1. Introduction;51
8.2;2. Early Concepts, Misconceptions, Unsettled Battles, and Persistent Confusion;52
8.3;3. The Discovery of "Natural" Free Radicals ;55
8.4;4. Love Affairs Between Hydroperoxides and Thiolates or Selenolates;58
8.4.1;4.1. Glutathione peroxidases;58
8.4.2;4.2. Peroxiredoxins;63
8.4.3;4.3. Other thiol peroxidases;66
8.4.4;4.4. Redoxins;66
8.5;5. Toward Regulatory Circuits with Puzzle Stones from Redox Biochemistry;68
8.5.1;5.1. Triggering signals;69
8.5.2;5.2. Hydroperoxide sensors;70
8.5.3;5.3. Signal transducers;72
8.5.4;5.4. Targets;73
8.5.5;5.5. Shut-off switches and restoration of starting conditions;73
8.5.6;5.6. Signaling versus defense or modulation of redox signaling by competition;75
8.6;6. Conclusions and Perspectives;76
8.7;References;77
9;Chapter 2: Mass Spectrometry-Based Methods for the Determination of Sulfur and Related Metabolite Concentrations in Cell Extracts;90
9.1;1. Introduction;91
9.2;2. Analytical Methods for the Determination of Sulfur and Amino Acid Metabolites;93
9.3;3. Procedures;95
9.3.1;3.1. Cell growth and 15N metabolic labeling;95
9.3.2;3.2. Metabolite extraction;97
9.4;4. Absolute LC-ESI-MS/MS Quantification of Thiol and Amino Acid Metabolites in Yeast Extracts;98
9.4.1;4.1. Sample preparation;98
9.4.2;4.2. Liquid chromatography;99
9.4.3;4.3. Mass spectrometry;101
9.4.4;4.4. Data processing;103
9.4.5;4.5. Method validation;106
9.5;5. Qualitative and Quantitative Determination of Thiol and Amino Acid Metabolites in Yeast Extracts by Using an LTQ-Orbitrap Mass Spectrometer;113
9.5.1;5.1. Sample preparation;113
9.5.2;5.2. Direct introduction;113
9.5.3;5.3. LC/MS;113
9.5.4;5.4. Data processing;115
9.6;6. Discussion;117
9.7;7. Summary;119
9.8;Acknowledgments;122
9.9;References;122
10;Chapter 3: Use of Dimedone-Based Chemical Probes for Sulfenic Acid Detection: Evaluation of Conditions Affecting Probe Incorporation into Redox-Sensitive Proteins;126
10.1;1. Introduction;127
10.2;2. Materials;130
10.2.1;2.1. Solutions;130
10.2.2;2.2. Chemical modification agents;131
10.2.3;2.3. Proteins;132
10.3;3. Methods;132
10.3.1;3.1. Characterization of "DCP"-linked compounds;132
10.3.2;3.2. Protocols for labeling cysteine sulfenic acids within cellular proteins;135
10.4;4. Summary;141
10.5;Acknowledgments;141
10.6;References;142
11;Chapter 4: Use of Dimedone-Based Chemical Probes for Sulfenic Acid Detection: Methods to Visualize and Identify Labeled Proteins;144
11.1;1. Introduction;145
11.2;2. Biotin-Based Affinity Capture to Identify Proteins Containing Cysteine Sulfenic Acids;147
11.2.1;2.1. Materials;148
11.2.2;2.2. Methods;149
11.3;3. Detection Methods to Identify Oxidized Proteins and Cysteines;152
11.3.1;3.1. Targeted approaches: Western blot of affinity-enriched proteins to analyze proteins of interest;152
11.3.2;3.2. Targeted approaches: Immunoprecipitation of protein of interest followed by Western blot to detect biotin;153
11.3.3;3.3. Controls for endogenous biotinylation;155
11.3.4;3.4. Global approaches: Identification of overall sulfenic acid levels for cellular proteins in response to stimuli;155
11.3.5;3.5. Global approaches: Identification of oxidized proteins by mass spectrometry after biotin affinity capture;159
11.3.6;3.6. Identification of oxidized cysteine by MS–MS analysis;159
11.4;4. Summary;161
11.5;Acknowledgments;162
11.6;References;162
12;Chapter 5: Formation and Reactions of Sulfenic Acid in Human Serum Albumin;166
12.1;1. Introduction;167
12.2;2. Preparation of Albumin Solutions;168
12.2.1;2.1. Source of albumin for biochemical studies;168
12.2.2;2.2. Albumin delipidation;168
12.2.3;2.3. Albumin thiol reduction;169
12.2.4;2.4. Albumin quantification;169
12.2.5;2.5. Thiol quantification;169
12.2.6;2.6. Thiol blockage;172
12.3;3. Preparation of Oxidized Albumin;172
12.4;4. Detection of Albumin Sulfenic Acid;174
12.4.1;4.1. Sodium arsenite;175
12.4.2;4.2. 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl);175
12.4.3;4.3. Dimedone and mass spectrometry;176
12.4.4;4.4. Glutathione;179
12.5;5. Quantification of Albumin Sulfenic Acid Using Thionitrobenzoate (TNB);180
12.6;6. Reactivity of Sulfenic Acid;181
12.7;7. Detection of Albumin Sulfinic Acid;183
12.8;8. Conclusions;183
12.9;Acknowledgments;183
12.10;References;184
13;Chapter 6: Determination of GSH, GSSG, and GSNO Using HPLC with Electrochemical Detection;186
13.1;1. Introduction;187
13.2;2. Methods;188
13.2.1;2.1. High-performance liquid chromatography with electrochemical detection;188
13.2.2;2.2. Hydrodynamic voltammogram of GSH, GSNO, and GSSG;189
13.2.3;2.3. GSNO detection in biological samples: Effect of sample preparation;191
13.2.4;2.4. Measurement of GSNO reductase activity using HPLC;193
13.3;3. Summary;194
13.4;Acknowledgments;195
13.5;References;195
14;Chapter 7: Measurement of Mixed Disulfides Including Glutathionylated Proteins;198
14.1;1. Introduction;199
14.2;2. Chemical Quantification of PSSX, PSSG, and PSSC;200
14.2.1;2.1. Principle;200
14.2.2;2.2. Reagents and solutions;201
14.2.3;2.3. Experimental procedure;201
14.2.4;2.4. Results;203
14.2.5;2.5. Typical data;203
14.2.6;2.6. Importance of basic pH;204
14.2.7;2.7. Importance of DTT and PSH;204
14.3;3. Visualization of PSSG by Western Blot;205
14.3.1;3.1. Principle;205
14.3.2;3.2. Methods and procedures;205
14.3.3;3.3. Typical results;206
14.4;Acknowledgments;206
14.5;References;207
15;Chapter 8: Detection and Quantification of Protein Disulfides in Biological Tissues: A Fluorescence-BasedProteomic Approach;210
15.1;1. Introduction;212
15.2;2. Material and Methods;213
15.2.1;2.1. Animals;213
15.2.2;2.2. Gel electrophoresis-based protein disulfide assay;213
15.2.3;2.3. Selection of the alkylating agents for measuring protein disulfide levels;214
15.2.4;2.4. Transformation of fluorescence units to nmoles of protein disulfide;216
15.2.5;2.5. 2D gel electrophoresis;216
15.2.6;2.6. Identification of proteins by MALDI-TOF/MS;217
15.3;3. Results;218
15.3.1;3.1. Measurement of changes in protein disulfide levels in response to oxidative stress;218
15.3.2;3.2. Measurement of changes in protein disulfide levels in young and old mice;218
15.3.3;3.3. Changes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity in young and old mice;220
15.4;4. Discussion;223
15.5;Acknowledgments;224
15.6;References;224
16;Chapter 9: Measurement and Identification of S-Glutathiolated Proteins;228
16.1;1. Introduction;229
16.2;2. Chemical Methods for the Measurement of Glutathiolated Proteins;231
16.2.1;2.1. Release of protein-bound GSH by oxidation;232
16.3;3. Detection of Glutathiolated Proteins by ESI/MS;233
16.3.1;3.1. Mass spectrometric analysis;234
16.4;4. Identification of S-Glutathiolated Proteins in Cells and Tissues;235
16.4.1;4.1. Western blot analysis;235
16.4.2;4.2. Immunohistochemical staining;240
16.4.3;4.3. Radioactive tagging;241
16.4.4;4.4. Biotin labeling;242
16.5;5 Conclusions;243
16.6;Acknowledgment;245
16.7;References;245
17;Chapter 10: Proteome Screens for Cys Residues Oxidation: The Redoxome;248
17.1;1. Introduction;249
17.2;2. General Considerations;250
17.2.1;2.1. Limits in the access to Cys-residues redox modifications;250
17.2.2;2.2. Acid quenching and Cys differential labeling;250
17.3;3. Overview of the Different Methods;251
17.3.1;3.1. 2DE-based methods;251
17.3.2;3.2. Shotgun proteomic: The MS-based ICAT technology;254
17.4;4. Results and Discussion;255
17.4.1;4.1. Methods;255
17.4.2;4.2. Results;259
17.5;5. Conclusions;263
17.6;Acknowledgments;263
17.7;References;263
18;Chapter 11: Identification by MS/MS of Disulfides Produced by a Functional Redox Transition;266
18.1;1. Introduction;267
18.2;2. MS/MS Identification of Redox-Switches in Protein;268
18.3;3. Analytical Procedure;269
18.3.1;3.1. Protein reduction and oxidation;269
18.3.2;3.2. Enzymatic digestions;270
18.3.3;3.3. LC–ESI-MS/MS;270
18.3.4;3.4. Data handling of mass spectra;270
18.3.5;3.5. Identification of the disulfide;270
18.4;4. Discussion;271
18.5;References;273
19;Chapter 12: Mass Spectrometry Approaches for the Redox Characterization of Protein Cysteine Residues: The Case of theTranscription Factor Pax-8;276
19.1;1. Introduction;277
19.2;2. Materials and Methods;279
19.2.1;2.1. Protein expression and functional analysis;279
19.2.2;2.2. Alkylation of protein samples with iodoacetamide and ESI-quadrupole-MS analysis ;281
19.2.3;2.3. Enzymatic digestion and MALDI-TOF peptide-mapping experiments;281
19.3;3. Results;281
19.3.1;3.1. Combined functional and MS analysis of the Pax-8 Prd domain under various redox conditions;281
19.3.2;3.2. MS assignment of modified Cys residues in Pax-8 Prd domain;285
19.3.3;3.3. Functional role of the modified Cys residues on Pax-8 activity;288
19.4;4. Conclusions and Future Perspectives;290
19.5;Acknowledgments;292
19.6;References;292
20;Chapter 13: A Simple Method to Systematically Study Oxidatively Modified Proteins in Biological Samples and Its Applications;300
20.1;1. Introduction;301
20.2;2. Materials;303
20.2.1;2.1. Chemicals and other materials;303
20.3;3. Methods;304
20.4;4. Discussion;306
20.4.1;4.1. Advantages of the simple redox-based Cys-targeted proteomics method;306
20.4.2;4.2. Limitations of the redox-based Cys-targeted proteomics and alternative approaches;309
20.5;Acknowledgment;311
20.6;References;311
21;Chapter 14: Direct and Indirect Detection Methods for the Analysis of S-Nitrosylated Peptides and Proteins;314
21.1;1. Introduction;315
21.2;2. Indirect Detection of S-Nitrosylated Proteins: His-tag Switch;317
21.2.1;2.1. Protocol for the analysis of S-nitrosylation using the His-tag switch;318
21.3;3. Direct Analysis of S-Nitrosylation by MS;323
21.3.1;3.1. Protocol for the direct analysis of S-nitrosylated peptides;323
21.4;4. Conclusions;327
21.5;References;327
22;Chapter 15: A Rapid Approach for the Detection, Quantification, and Discovery of Novel Sulfenic Acid or S-Nitrosothiol Modified Proteins Using a Biotin-Switch Method;330
22.1;1. Introduction;331
22.1.1;1.1. Nitrosative protein oxidation;332
22.1.2;1.2. Sulfenic acid formation;337
22.2;2. Overview of Analytical Strategy;341
22.2.1;2.1. Generating S-nitrosocysteine;342
22.2.2;2.2. Tissue preparation;342
22.2.3;2.3. Detection of modified proteins using the biotin-switch method;343
22.2.4;2.4. Detection and purification of modified proteins after the biotin-switch method;343
22.3;3. Conclusions;345
22.4;Acknowledgments;346
22.5;References;346
23;Chapter 16: Protein Adducts of Aldehydic Lipid Peroxidation Products: Identification and Characterizationof Protein Adducts Using anAldehyde/Keto-Reactive Probein Combination with MassSpectrometry;354
23.1;1. Introduction;355
23.2;2. Modification of Proteins by Aldehydic Lipid Peroxidation Products;356
23.3;3. Redox Proteomics of Protein Targets of Reactive Lipid Peroxidation Products;357
23.4;4. Mass Spectrometry-Based Approaches for the Identification and Characterization of Protein Adducts of Aldehydic Lipid Peroxidation Products;359
23.5;5. Experimental Strategy of Using an Aldehyde/Keto-Reactive Probe for the Targeted Analysis of Protein Adducts of Aldehydic Lipid Peroxidation Products;360
23.5.1;5.1. ARP-labeling of aldehydic protein adducts of 2-alkenals and tryptic proteolysis;361
23.5.2;5.2. Enrichment of ARP-labeled peptide adducts using biotin avidin affinity chromatography;362
23.5.3;5.3. Tandem mass spectrometry for peptide identification and determining the site of adduction;363
23.5.4;5.4. Searching of MS/MS data against protein sequence databases for the identification of peptide sequences and peptide adducts;365
23.6;6. Applications of the ARP-Labeling Strategy;365
23.7;7. Interpretation of MS/MS Spectra of Protein Adducts and the Use of Diagnostic Marker Ions;372
23.8;8. Conclusion;373
23.9;Acknowledgment;376
23.10;References;377
24;Author Index;380
25;Subject Index;402
26;Color Plate;410