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電子書籍詳細


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質量分析に基づくケミカルプロテオミクス

Mass Spectrometry-Based Chemical Proteomics

1

Tao, W. Andy (EDT)   Zhang, Ying (EDT)

Wiley 2019/07
448p.
出版国: US
ISBN: 9781118969557
eISBN: 9781118970201
KNPID: EY00355878
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Full Description

PROVIDES STRATEGIES AND CONCEPTS FOR UNDERSTANDING CHEMICAL PROTEOMICS, AND ANALYZING PROTEIN FUNCTIONS, MODIFICATIONS, AND INTERACTIONS—EMPHASIZING MASS SPECTROMETRY THROUGHOUT

Covering mass spectrometry for chemical proteomics, this book helps readers understand analytical strategies behind protein functions, their modifications and interactions, and applications in drug discovery. It provides a basic overview and presents concepts in chemical proteomics through three angles: Strategies, Technical Advances, and Applications. Chapters cover those many technical advances and applications in drug discovery, from target identification to validation and potential treatments.

The first section of Mass Spectrometry-Based Chemical Proteomics starts by reviewing basic methods and recent advances in mass spectrometry for proteomics, including shotgun proteomics, quantitative proteomics, and data analyses. The next section covers a variety of techniques and strategies coupling chemical probes to MS-based proteomics to provide functional insights into the proteome. In the last section, it focuses on using chemical strategies to study protein post-translational modifications and high-order structures.

  • Summarizes chemical proteomics, up-to-date concepts, analysis, and target validation
  • Covers fundamentals and strategies, including the profiling of enzyme activities and protein-drug interactions
  • Explains technical advances in the field and describes on shotgun proteomics, quantitative proteomics, and corresponding methods of software and database usage for proteomics
  • Includes a wide variety of applications in drug discovery, from kinase inhibitors and intracellular drug targets to the chemoproteomics analysis of natural products
  • Addresses an important tool in small molecule drug discovery, appealing to both academia and the pharmaceutical industry

Mass Spectrometry-Based Chemical Proteomics is an excellent source of information for readers in both academia and industry in a variety of fields, including pharmaceutical sciences, drug discovery, molecular biology, bioinformatics, and analytical sciences.

Table of Contents

Preface xv

1 Protein Analysis by Shotgun Proteomics 1
Yu Gao and John R. Yates III

1.1 Introduction 1

1.1.1 Terminology 1

1.1.2 Power of Shotgun Proteomics 1

1.1.3 Advantage of Shotgun Proteomics 2

1.2 Overview of Shotgun Proteomics 2

1.3 Sample Preparation 4

1.3.1 Protein Separation 4

1.3.1.1 Overview 4

1.3.1.2 2D‐Gel Approach 4

1.3.1.3 Separation of Membrane Protein 5

1.3.1.4 Subcellular Fractionation 5

1.3.1.5 Protein Enrichment 6

1.3.1.6 Phosphoprotein 6

1.3.1.7 Glycoprotein 6

1.3.1.8 AP–MS and Interactome 7

1.3.2 Protein Modification 8

1.3.2.1 Overview 8

1.3.2.2 Reduction of Disulfide Bond and Alkylation 8

1.3.2.3 Chemical Crosslinking 8

1.3.2.4 Proximity Labeling 9

1.3.3 Protein Digestion 9

1.4 Peptide Separation and Data Acquisition 11

1.4.1 Peptide Separation 11

1.4.1.1 Reversed Phase (RP) 11

1.4.1.2 HILIC 11

1.4.1.3 MudPIT 11

1.4.1.4 Capillary Electrophoresis 13

1.4.2 Peptide Ionization 13

1.4.3 Mass Analyzer 13

1.4.4 Peptide Fragmentation Method 15

1.4.4.1 CID/HCD 15

1.4.4.2 ETD/ECD 16

1.4.4.3 IRMPD/UVPD 16

1.4.5 Acquisition Mode 17

1.5 Informatics 17

1.5.1 Peptide Identification 18

1.5.1.1 Database Search 18

1.5.1.2 Spectral Library Search 21

1.5.1.3 De novo Sequencing 22

1.5.1.4 Peptide‐Centric Analysis 23

1.5.2 Peptide/Protein Quantitation 23

1.5.2.1 Labeled Quantitation 23

1.5.2.2 Label‐Free Quantitation 27

1.5.3 Protein Inference 29

References 31

2 Quantitative Proteomics for Analyses of Multiple Samples in Parallel with Chemical Perturbation 39
Amanda Rae Buchberger, Jillian Johnson, and Lingjun Li

2.1 Introduction 39

2.2 Relative and Absolute Label‐Free Quantitation Strategies 40

2.3 Stable Isotope‐Based Quantitative Proteomics 42

2.3.1 Relative Quantitation 42

2.3.2 Absolute Quantitation 47

2.4 Conclusion 48

2.5 Methodology 50

2.6 Notes 52

Acknowledgments 55

References 56

3 Chemoproteomic Analyses by Activity‐Based Protein Profiling 67
Bryan J. Killinger, Kristoffer R. Brandvold, Susan J. Ramos‐Hunter, and Aaron T. Wright

3.1 Introduction 67

3.2 How ABPP Works 68

3.3 ABPP Probe Design 71

3.3.1 Mechanism‐Based Probes 72

3.3.2 Reactivity‐Based Probes 74

3.3.3 Photoaffinity Probes 74

3.4 ABPP and Mass Spectrometry for Chemoproteomics 75

3.4.1 Determining ABP Target Identity 75

3.4.2 Considerations for Analyzing ABP Targets with MS 77

3.4.3 Determining the Site of ABP Labeling 78

3.4.4 Quantification of ABPP Probe Targets 80

3.4.4.1 Label‐Free Methods 80

3.4.4.2 Isotopic Methods 81

3.5 ABPP Applications and Recent Advances 83

3.5.1 Using ABPs for Functional Protein Annotation 83

3.5.2 ABPPs Applied to Microbes and Their Communities 84

3.6 ABPP Applied to Drug Discovery 88

3.7 Comparative, Competitive, and Convolution ABPP 90

3.8 Conclusions and The Outlook of ABPP 91

Acknowledgements 91

References 91

4 Activity‐Based Probes for Profiling Protein Activities 101
Kasi V. Ruddraraju and Zhong‐Yin Zhang

4.1 Introduction 101

4.2 Design of Activity‐Based Probes 102

4.2.1 The Reactive Group 102

4.2.2 The Linker 104

4.2.3 The Tag 104

4.3 Analytical Platforms for ABPP 105

4.3.1 Gel‐Based Platforms 105

4.3.2 Mass Spectrometry Platforms for ABPP 106

4.3.3 Microarray Platform for ABPP 107

4.3.4 Capillary Electrophoresis Platform for ABPP 107

4.4 Classes of Enzymes Studied by ABPP 108

4.4.1 Serine Hydrolases 108

4.4.2 Cysteine Proteases 109

4.4.3 Metallohydrolases 110

4.4.4 Glycosidases 111

4.4.5 Protein Kinases 114

4.4.6 Protein Phosphatases 116

4.5 Conclusions 119

Acknowledgment 120

References 120

5 Chemical Probes for Proteins and Networks 127
Scott Lovell, Charlotte L. Sutherell, and Edward W. Tate

5.1 Introduction 127

5.1.1 Probe Design and Validation 128

5.1.2 Application to a Proteomics Workflow 129

5.1.3 Quantitative Chemical Proteomics 131

5.2 Application of Metabolic Chemical Probes to Lipidated Protein Networks 132

5.2.1 Chemical Probes for N‐Myristoylation 133

5.2.2 Chemical Probes for Hedgehog Proteins 136

5.3 Chemical Probes for Target Identification 137

5.3.1 Identifying New Target Profiles of Sulforaphane in Breast Cancer Cells 138

5.3.2 Target Profiling of Zerumbone Using a Novel Clickable Probe 140

5.4 Protocol 143

5.4.1 Introduction 143

5.4.2 Materials 143

5.4.2.1 Chemical Tools 143

5.4.2.2 Cell Culture 143

5.4.2.3 Cell Lysis, Enrichment and Sample Preparation 144

5.4.2.4 Click Chemistry and Enrichment 144

5.4.2.5 Proteomics Sample Preparation 144

5.4.2.6 Proteomics Analysis 144

5.4.3 Method 144

5.4.3.1 HeLa Cell Culture and Preparation of Spike‐in Standard 144

5.4.3.2 Preparation of Cell Lysates for Protein Enrichment 145

5.4.3.3 Pull‐Down Experiments and Sample Preparation 145

5.4.3.4 LC–MS/MS Analysis 147

5.4.3.5 Data Analysis 147

5.4.3.6 Identification of N‐Terminal Myristoylated Peptides 151

5.5 Notes 152

References 153

6 Probing Biological Activities with Peptide and Peptidomimetic Biosensors 159
Laura J. Marholz, Tzu-Yi Yang, and Laurie L. Parker

6.1 Introduction 159

6.2 Peptide Biosensors for Assignment and Characterization of Enzymatic Reactions and Substrate Specificity 160

6.3 Screening Inhibitors and Detecting Ligand Interactions 165

6.4 Diagnostic and Clinical Applications 168

6.5 Profiling Enzymatic Activity 172

6.6 Protocol 178

Materials 179

Methods 180

6.7 Conclusion 182

References 182

7 Chemoselective Tagging to Promote Natural Product Discovery 187
Emily J. Tollefson and Erin E. Carlson

7.1 Introduction 187

7.2 Nonreversible Mass Spectrometry Tags 189

7.2.1 Azides and Alkynes 189

7.2.2 Thiols 192

7.2.3 Aminooxy 194

7.3 Reversible Enrichment Tags 195

7.3.1 Boronic Acids 195

7.3.2 Hydrazines 196

7.3.3 Silanes 196

7.3.4 Disulfides 197

7.4 Conclusions 198

7.5 Protocol for Enrichment of Carboxylic‐Acid‐Containing Natural Products 198

7.5.1 Dialkylsiloxane Resin Synthesis 198

7.5.2 Production of S. rochei Extract 200

7.5.3 Chemoselective Capture 200

7.5.4 Release of Carboxylic‐Acid‐Containing Compounds from Resin 201

References 201

8 Identification and Quantification of Newly Synthesized Proteins Using Mass‐Spectrometry Based Chemical Proteomics 207
Suttipong Suttapitugsakul, Haopeng Xiao, and Ronghu Wu

8.1 Introduction 207

8.2 Protein Labeling to Study Newly Synthesized Proteins 209

8.2.1 Radioactive Labeling 209

8.2.2 Protein Labeling with Fluorescent Probes 209

8.2.3 SILAC Labeling 210

8.2.4 Protein Labeling with Noncanonical Amino Acids 210

8.3 Global Identification of Newly Synthesized Proteins by Noncanonical Amino Acids and MS 212

8.4 Comprehensive Quantification of Newly Synthesized Proteins by MS 213

8.5 Materials 217

8.5.1 Cell Culture and AHA Labeling 217

8.5.2 Cell Lysis 218

8.5.3 Enrichment of Newly Synthesized Proteins Using Click Chemistry 218

8.5.4 On‐Bead Protein Reduction, Alkylation, and Digestion 218

8.5.5 Peptide Desalting 218

8.5.6 TMT Labeling 219

8.5.7 Peptide Fractionation 219

8.5.8 StageTips 219

8.5.9 LC–MS/MS Analysis 219

8.5.10 Database Searches and Data Filtering 220

8.6 Methods 220

8.6.1 Cell Culture with AHA Labeling 220

8.6.2 Cell Lysis and Protein Extraction 220

8.6.3 Enrichment of Newly Synthesized Proteins 220

8.6.4 On‐Bead Reduction, Alkylation, and Digestion 221

8.6.5 Peptide Desalting 221

8.6.6 TMT Labeling 222

8.6.7 Peptide Fractionation 222

8.6.8 StageTip Purification 222

8.6.9 LC–MS/MS Analysis 223

8.6.10 Database Searches, Data Filtering, and Half‐Life Calculation of Newly Synthesized Proteins 223

Acknowledgements 224

References 224

9 Tracing Endocytosis by Mass Spectrometry 231
Mayank Srivastava, Ying Zhang, Linna Wang, and W. Andy Tao

9.1 Introduction 231

9.2 Clathrin‐Mediated Endocytosis 232

9.2.1 Proteins Involved in the Formation of Clathrin‐Coated Vesicles 233

9.2.2 Molecular Mechanism for CCV Formation 234

9.2.3 Vesicle Uncoating and Fusion with Endosomal Compartments 237

9.3 Mass Spectrometry as a Tool to Study Endocytosis 237

9.3.1 Isolation of Clathrin‐Coated Vesicles and Analysis Using Mass Spectrometry 238

9.3.2 Chemical Proteomic Approaches for Studying the Endocytosis 240

9.3.2.1 Identification of Receptor by Ligand‐based–Receptor Capture (LRC) Technology 240

9.3.2.2 Studying the Entry and Trafficking of Nanoparticles Using Time‐Resolved Chemical Proteomic Approach 241

9.4 Protocols for TITAN 243

9.4.1 Materials 243

9.4.2 Dendrimer Functionalization 245

9.4.2.1 Synthesis of Masked Aldehyde Handle 245

9.4.2.2 Functionalization of Dendrimer 245

9.4.3 Internalization of Dendrimer by HeLa and MS Sample Preparation 247

9.4.4 Mass Spectrometry and Data Analysis 249

9.5 Conclusion and Future Directions 250

References 251

10 Functional Identification of Target by Expression Proteomics (FITExP) 257
Massimiliano Gaetani and Roman A. Zubarev

10.1 Introduction 257

10.2 FITExP Protocol 261

10.2.1 Cell Line(s) and Drugs/Compounds Selection 261

10.2.2 Drug Treatments of Cell Cultures 261

10.2.3 Cell Lysis and Protein Extraction 262

10.2.4 Estimation of Protein Concentration and Protein Sample Processing 263

10.2.5 Protein Digestion 263

10.2.6 Peptide TMT (Tandem Mass Tag) Labeling and Desalting 263

10.2.7 High pH Fractionation TMT 264

10.2.8 Mass Spectrometry Analysis 264

10.2.9 Data Analysis 265

References 265

11 Target Discovery Using Thermal Proteome Profiling 267
Sindhuja Sridharan, Ina Günthner, Isabelle Becher, Mikhail Savitski, and Marcus Bantscheff

11.1 Introduction 267

11.2 Thermodynamics of Ligand Binding as a Measure of Target Engagement 270

11.3 Thermal Proteome Profiling – Proteome‐wide Detection of Drug–Target Interactions 273

11.3.1 Overview 273

11.3.2 Distinguishing Direct Drug Targets from Downstream Effectors of Drug Action 273

11.4 Experimental Formats 275

11.4.1 Temperature‐Range Experiment (TPP‐TR) 275

11.4.2 Compound Concentration‐Range Experiment (TPP‐CCR) 277

11.4.3 Two‐Dimensional TPP (2D‐TPP) 278

11.5 Experimental Protocol 278

11.6 Reagents 280

11.6.1 Step 1: Compound Treatment 280

11.6.2 Step 2: Temperature Treatment 281

11.6.3 Step 3: Protein Digestion and Labeling 282

11.6.4 Step 4: Mass Spectrometric Analysis of Samples 283

11.6.5 Step 5: Peptide and Protein Identification and Quantification 283

11.6.6 Step 6: Data Handling and Analysis 284

11.7 Present Challenges with TPP 284

11.8 CETSA to TPP – Where are We Heading? 285

References 287

12 Chemical Strategies to Glycoprotein Analysis 293
Joseph L. Mertz, Christian Toonstra, and Hui Zhang

12.1 Introduction 293

12.2 Sample Preparation Strategies for Glycoproteomics 297

12.2.1 Enzymatic/Chemical Modification for Glycopeptide Enrichment 297

12.2.2 Enrichment of Glycans or Glycopeptides by Physical–Chemical Approaches 300

12.3 MS Analysis 302

12.3.1 Glycoproteomic Analysis by Mass Spectrometry 302

12.3.2 Bioinformatics and Data Analysis 304

12.4 Conclusions 306

References 307

13 Proteomic Analysis of Protein–Lipid Modifications: Significance and Application 317
Kiall F. Suazo, Garrett Schey, Chad Schaber, Audrey R. Odom John, and Mark D. Distefano

13.1 Introduction 317

13.2 Chemical Proteomic Approach to Identify Lipidated Proteins 318

13.2.1 Fatty Acylation 322

13.2.1.1 N‐Myristoylation 323

13.2.1.2 S‐Palmitoylation 325

13.2.2 Prenylation 328

13.2.3 Modification with Cholesterol and GPI Anchors 330

13.3 Protocol for Proteomic Analysis of Prenylated Proteins 331

13.3.1 Materials 332

13.3.1.1 Reagents 332

13.3.1.2 Equipment 333

13.3.1.3 Reagents and Instrument Setup 333

13.3.2 Procedure 334

13.3.2.1 Labeling with Probe 334

13.3.2.2 Isolating Parasites via Saponin Lysis 335

13.3.2.3 In‐gel Fluorescence Analysis 335

13.3.2.4 Biotinylation and Streptavidin Pull‐down 336

13.3.2.5 Sample Preparation for LC–MS/MS Analysis 337

13.3.2.6 LC–MS/MS Analysis 337

13.3.2.7 Proteomic Data Analysis Using Spectral Counting 338

13.3.3 Results 338

References 341

14 Site‐Specific Characterization of Asp‐ and Glu‐ADP‐Ribosylation by Quantitative Mass Spectrometry 349
Shuai Wang, Yajie Zhang, and Yonghao Yu

14.1 Introduction 349

14.2 Materials 353

14.2.1 Cell Culture 353

14.2.2 Generation of Stable Cell Lines Expressing shPARG 353

14.2.3 Sample Preparation for Mass Spectrometry 353

14.2.4 Mass Spectrometry Analysis 354

14.2.5 Equipment 354

14.3 Methods 354

14.3.1 Generation of shPARG‐Expressing Cell Line 354

14.3.2 SILAC Cell Culture 355

14.3.3 Cell Lysis 355

14.3.4 Reduction, Alkylation, and Precipitation of Proteins 355

14.3.5 Protein Digestion and Enrichment of the PARylated Peptides 356

14.3.6 Cleanup of the Peptide 357

14.3.7 Mass Spectrometry Analysis and Data Processing 357

14.4 Notes 357

Acknowledgements 358

References 358

15 MS‐Based Hydroxyl Radical Footprinting: Methodology and Application of Fast Photochemical Oxidation of Proteins (FPOP) 363
Ben Niu and Michael L. Gross

15.1 Introduction 363

15.1.1 General Approaches for Mapping Protein Conformations 363

15.1.2 MS‐Based Approaches 364

15.2 Generation of Hydroxyl Radicals 365

15.2.1 Fenton and Fenton‐like Chemistry 365

15.2.2 Electron-Pulse Radiolysis 368

15.2.3 High‐Voltage Electrical Discharge 370

15.2.4 Synchrotron X‐ray Radiolysis of Water 371

15.2.5 Plasma Formation of OH Radicals 372

15.2.6 Photolysis of Hydrogen Peroxide 374

15.3 Fast Photochemical Oxidation of Proteins (FPOP) 375

15.3.1 FPOP Footprints Faster than Protein Folding/Unfolding 377

15.3.2 FPOP Dosimetry 378

15.3.3 Primary Radical Lifetime and Adjustment of Radical Scavengers 379

15.3.4 Radical Lifetimes Can Be Milliseconds 381

15.3.5 Differential Scavenging and Use of a Reporter Peptide in FPOP 381

15.3.6 New Reactive Reagents for the FPOP Platform 383

15.4 Applications of FPOP 384

15.4.1 FPOP for Protein–Protein Interactions and Epitope Mapping 384

15.4.2 FPOP for Protein Aggregation/Oligomerization 387

15.4.3 FPOP for Protein Dynamics 390

15.4.4 FPOP for Protein Folding 391

15.4.5 FPOP for Characterizing Membrane Proteins 394

15.5 Conclusions 395

References 396

Index 417