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Book Details
Abstract
Glycans play a vital role in modulating protein structure and function from involvement in protein folding, solubility and stability to regulation of tissue distribution, recognition specificity, and biological activity. They can act as both positive and negative regulators of protein function, providing an additional level of control with respect to genetic and environmental conditions.
Due to the complexity of glycosylated protein forms, elucidating structural and functional information has been challenging task for researchers but recent development of chemical biology-based tools and techniques is bridging these knowledge gaps. This book provides a thorough review of the current state of glycoprotein chemical biology, describing the development and application of glycoprotein and glycan synthesis technologies for understanding and manipulating protein glycosylation.
Table of Contents
| Section Title | Page | Action | Price |
|---|---|---|---|
| Cover | Cover | ||
| Chemical Biology of Glycoproteins | i | ||
| Preface | v | ||
| Dedication | ix | ||
| Contents | xi | ||
| Chapter 1 - Introduction: General Aspects of the Chemical Biology of Glycoproteins | 1 | ||
| 1.1 Introduction | 1 | ||
| 1.1.1 Complexity of Protein Glycosylation | 2 | ||
| 1.1.2 Strategies and Methods to Study Protein Glycosylation | 3 | ||
| 1.2 Chemical Biology of Glycoproteins | 5 | ||
| 1.2.1 Types of Protein Glycosylation | 5 | ||
| 1.2.2 Biosynthesis of Glycoproteins | 7 | ||
| 1.2.3 Structural and Functional Consequences of Protein Glycosylation | 8 | ||
| 1.2.4 Methods to Prepare Homogeneous Glycopeptides and Glycoproteins | 10 | ||
| 1.2.5 Chemical Glycobiology and Applications | 11 | ||
| 1.3 Conclusion | 12 | ||
| References | 13 | ||
| Chapter 2 - Chemical Biology of Protein N-Glycosylation | 20 | ||
| 2.1 Introduction | 20 | ||
| 2.2 Biosynthesis and Intracellular Functions of N-Glycans of Glycoproteins | 21 | ||
| 2.3 Inhibitors of Glycan-Processing Enzymes for Controlling Protein N-Glycosylation | 23 | ||
| 2.4 Global Metabolic Enzyme Inhibitors for Perturbing Protein N-Glycosylation | 25 | ||
| 2.5 Metabolic Glycoengineering of Cell-Surface Glycoproteins | 28 | ||
| 2.6 Chemoenzymatic Synthesis and Glycosylation Remodeling Toward Homogeneous N-Glycoproteins | 31 | ||
| 2.6.1 Generation of Novel ENGase-Based Glycosynthases for N-Glycosylation Remodeling and N-Glycoprotein Synthesis | 34 | ||
| 2.6.2 Chemoenzymatic Fc Glycan Remodeling of Therapeutic Monoclonal Antibodies | 37 | ||
| 2.6.3 Combined E. coli Expression and Chemoenzymatic Glycan Remodeling to Produce Humanized N-Glycoproteins | 39 | ||
| 2.7 Conclusion | 41 | ||
| Acknowledgements | 41 | ||
| References | 42 | ||
| Chapter 3 - Chemical Biology of Protein O-Glycosylation | 48 | ||
| 3.1 Introduction | 48 | ||
| 3.2 Biosynthesis of O-Glycoproteins | 51 | ||
| 3.2.1 α-O-GalNAc | 51 | ||
| 3.2.2 α-O-Man | 53 | ||
| 3.2.3 α-O-Fuc | 55 | ||
| 3.2.4 β-O-Glc | 56 | ||
| 3.2.5 β-O-GlcNAc | 56 | ||
| 3.3 Chemical Biology in Studying the Structural and Functional Consequences of Protein O-Glycosylation | 57 | ||
| 3.3.1 α-O-GalNAc | 57 | ||
| 3.3.1.1 Substrate Preferences of ppGalNAcTs | 59 | ||
| 3.3.1.2 Structural Effects of O-GalNAc Glycosylation | 60 | ||
| 3.3.1.3 Functional Effects of O-GalNAc Glycosylation | 63 | ||
| 3.3.2 α-O-Man | 64 | ||
| 3.3.2.1 Biosynthetic Pathway of O-Man Glycans | 65 | ||
| 3.3.2.2 Biophysical and Biological Effects of O-Mannosylation | 70 | ||
| 3.3.3 α-O-Fuc | 72 | ||
| 3.3.4 β-O-Glc | 75 | ||
| 3.3.5 β-O-GlcNAc | 76 | ||
| 3.4 Chemical Biology in Studying the Composition of Mixtures of O-Glycoproteins | 76 | ||
| 3.4.1 Glycoprotein Purification and Enrichment | 77 | ||
| 3.4.2 Glycoprotein Digestion and Glycopeptide Separation | 79 | ||
| 3.4.3 Glycopeptide Analysis | 80 | ||
| 3.4.4 Importance of Synthetic Glycopeptides in Protein Glycosylation Analysis | 81 | ||
| 3.5 Conclusion | 83 | ||
| References | 83 | ||
| Chapter 4 - Chemical Biology of O-GlcNAc Glycosylation | 94 | ||
| 4.1 Introduction | 94 | ||
| 4.2 Chemical Blockade of O-GlcNAc Addition | 100 | ||
| 4.2.1 Inhibitors of UDP–GlcNAc Biosynthesis | 100 | ||
| 4.2.2 Product Inhibition of OGT: UDP | 101 | ||
| 4.2.3 Alloxan | 102 | ||
| 4.2.4 Screening Approaches to Discover OGT Inhibitors | 103 | ||
| 4.2.5 Substrate Mimicry Approach to Discover OGT Inhibitors | 105 | ||
| 4.2.6 New Directions in OGT Inhibitor Development | 107 | ||
| 4.3 Chemical Blockade of O-GlcNAc Removal: OGA Inhibitors | 107 | ||
| 4.3.1 Streptozotocin | 107 | ||
| 4.3.2 PUGNAc | 108 | ||
| 4.3.3 NAG-Thiazoline | 110 | ||
| 4.3.4 GlcNAcstatins | 111 | ||
| 4.3.5 Thiamet-G | 112 | ||
| 4.3.6 Other Approaches to OGA Inhibition | 114 | ||
| 4.4 Detection of O-GlcNAcylated Substrates: Lectins and Antibodies | 114 | ||
| 4.5 Detection of O-GlcNAcylated Substrates: Mass Spectrometry-Based Methods | 116 | ||
| 4.5.1 Electron Transfer Dissociation MS | 117 | ||
| 4.5.2 MS Approaches to Complement ETD | 118 | ||
| 4.5.3 Chemical Derivatization of MS Samples for O-GlcNAc Enrichment and Detection | 119 | ||
| 4.6 Detection of O-GlcNAcylated Substrates: A Chemoenzymatic Approach | 121 | ||
| 4.7 Detection of O-GlcNAcylated Substrates: Metabolic Labeling Approaches | 125 | ||
| 4.7.1 N-Azidoacetylglucosamine (GlcNAz) | 125 | ||
| 4.7.2 N-Azidoacetylgalactosamine (GalNAz) | 127 | ||
| 4.7.3 N-Butynyl-Glucosamine (GlcNAlk) and Other Alkynylsugars | 129 | ||
| 4.7.4 Other Metabolic Labeling Reagents | 131 | ||
| 4.8 Methods to Deduce the Biochemical and Cellular Functions of O-GlcNAc | 132 | ||
| 4.8.1 Live-Cell Assays of OGT or OGA Activity | 132 | ||
| 4.8.2 Chemical Modification and Semi-Synthesis of Model Glycoproteins and Glycopeptides | 133 | ||
| 4.8.3 Photocrosslinking Tools to Capture O-GlcNAc-Mediated Protein–Protein Interactions | 135 | ||
| 4.9 Conclusions and Outlook | 137 | ||
| References | 137 | ||
| Chapter 5 - Chemical Synthesis and Engineering of N-Linked Glycoproteins | 150 | ||
| 5.1 Introduction | 150 | ||
| 5.2 Semisynthesis of N-Glycosylated Proteins | 152 | ||
| 5.2.1 Semisynthesis of N-Glycoprotein Mimics with Unnatural Glycosyl Linkages at Cysteines | 154 | ||
| 5.2.2 Semisynthesis of Glycoproteins with Natural N-Linkages on Asparagines | 156 | ||
| 5.3 Total Chemical Synthesis of N-Linked Glycoproteins | 165 | ||
| 5.3.1 Total Synthesis of N-Linked Glycoproteins Bearing N-Chitobioses | 167 | ||
| 5.3.2 Total Synthesis of N-Linked Glycoproteins Bearing “Wild-Type” N-Glycans | 167 | ||
| 5.3.3 Representative Synthetic Attempts Towards Producing N-Linked Glycoproteins with Multiple Disulfide Linkages | 172 | ||
| 5.4 Application of Chemically Synthesized N-Linked Glycoproteins to Biological Processes | 174 | ||
| 5.5 Conclusion | 183 | ||
| References | 183 | ||
| Chapter 6 - Chemoenzymatic Synthesis of N-Glycans | 188 | ||
| 6.1 Introduction | 188 | ||
| 6.2 Chemical Synthesis of Complex N-Glycans | 190 | ||
| 6.2.1 Assembly Strategy and Method of Glycosylation in Chemical Synthesis of N-Glycans | 190 | ||
| 6.2.2 Total Synthesis of Complex N-Glycans by Global Glycosylation of a Core Pentasaccharide | 191 | ||
| 6.2.3 Convergent Synthesis of Complex N-Glycans by Glycosylation of Core Trisaccharide | 191 | ||
| 6.3 Chemoenzymatic Synthesis of Complex N-Glycans | 194 | ||
| 6.3.1 A General Strategy for the Chemoenzymatic Synthesis of Asymmetrically Branched N-Glycans | 194 | ||
| 6.3.1.1 Chemical Synthesis of Asymmetric N-Glycans by Orthogonal Protection Strategy | 194 | ||
| 6.3.1.2 Enzymatic Extension to Synthesize Asymmetrically Branched N-Glycans | 196 | ||
| 6.3.2 Core Synthesis/Enzymatic Extension (CSEE) Strategy for Efficient Synthesis of N-Glycan Libraries | 196 | ||
| 6.3.2.1 Chemical Synthesis of N-Glycan Core Structures Through Convergent Block Coupling | 196 | ||
| 6.3.2.2 Enzymatic Extension to Obtain a N-Glycan Library | 199 | ||
| 6.4 Conclusion | 206 | ||
| References | 206 | ||
| Chapter 7 - Towards Synthesis of Heparan Sulfate Glycopeptides and Proteoglycans | 209 | ||
| 7.1 Structures, Biological Functions and Biosynthesis of Proteoglycans | 209 | ||
| 7.2 Chemical Synthesis of the PG Linker–Peptide Conjugates | 212 | ||
| 7.3 Synthesis of HS Glycopeptides | 224 | ||
| 7.4 Conclusion and Future Outlook | 229 | ||
| Acknowledgements | 229 | ||
| References | 229 | ||
| Chapter 8 - Chemoenzymatic Synthesis of Low-Molecular-Weight Heparin and Heparan Sulfate | 233 | ||
| 8.1 Introduction | 233 | ||
| 8.1.1 What are Heparan Sulfate, Heparin and Heparin-Derivatives | 233 | ||
| 8.1.2 Why are Synthetic Heparins and Heparan Sulfates Needed | 235 | ||
| 8.1.3 Types of Chemoenzymatic Synthesis | 236 | ||
| 8.2 Enzymes Required for Chemoenzymatic Synthesis | 238 | ||
| 8.2.1 Glycosyltransferases | 238 | ||
| 8.2.2 Sulfotransferases and C5-Epimerase | 239 | ||
| 8.3 Building Blocks Prepared for Chemoenzymatic Synthesis | 240 | ||
| 8.3.1 Acceptors | 240 | ||
| 8.3.2 Donors | 241 | ||
| 8.3.2.1 Natural UDP–Sugars | 241 | ||
| 8.3.2.2 Unnatural UDP–Sugars | 242 | ||
| 8.3.2.2.1 UDP–GlcNTFA.While most of the HS biosynthetic enzymes have been efficiently prepared via E. coli, only an active N-sulfotransfer... | 242 | ||
| 8.3.2.2.2\rOther Unnatural UDP–Sugars.The application of unnatural donors can also play an important role in synthesizing unnatural heparin... | 243 | ||
| 8.3.2.2.3\rChallenges for Unnatural Donors.Unnatural UDP–sugars have been successfully applied in synthesis of heparin analogs, establishin... | 243 | ||
| 8.3.3 Polysaccharide and Oligosaccharide Backbone | 243 | ||
| 8.4 Control of Product Through Sequential Enzymatic Modification | 244 | ||
| 8.5 Novel Chemoenzymatic Synthesis | 246 | ||
| 8.5.1 One-Pot Multienzyme System | 246 | ||
| 8.5.2 Fluorous-Tagging Techniques | 247 | ||
| 8.5.3 Solid-Phase Synthesis | 247 | ||
| 8.5.4 Immobilized Enzymes | 248 | ||
| 8.5.5 Immobilized Enzyme Cofactors | 248 | ||
| 8.6 Conclusion and Future Perspectives | 249 | ||
| References | 249 | ||
| Chapter 9 - Synthetic Studies of GPI-Anchored Peptides, Glycopeptides, and Proteins | 253 | ||
| 9.1 Introduction | 253 | ||
| 9.2 Biosynthesis of GPIs and GPI-Anchored Proteins | 255 | ||
| 9.2.1 Biosynthesis of GPI Anchors | 255 | ||
| 9.2.2 Posttranslational Attachment of GPIs to Proteins | 257 | ||
| 9.3 Synthesis of GPI-Anchored Proteins and Glycoproteins | 258 | ||
| 9.3.1 Chemical Total Synthesis of GPI-Anchored Peptides and Glycopeptides | 260 | ||
| 9.3.2 Synthesis of GPI-Anchored Peptides and Proteins via NCL | 262 | ||
| 9.3.3 Synthesis of GPI-Anchored Peptides, Glycopeptides, and Proteins via Enzymatic Ligation | 265 | ||
| 9.4 GPI-Anchored Proteomics Studies | 270 | ||
| 9.5 Concluding Remarks | 272 | ||
| Acknowledgements | 273 | ||
| References | 273 | ||
| Chapter 10 - Chemical Approaches to Image Protein Glycosylation | 282 | ||
| 10.1 Background | 282 | ||
| 10.2 Structure, Biosynthesis, and Function of Protein Glycans | 283 | ||
| 10.2.1 N-Linked Glycans | 284 | ||
| 10.2.2 Mucin-Type O-Linked Glycans | 284 | ||
| 10.2.3 Sialic Acids | 285 | ||
| 10.2.4 O-GlcNAc | 285 | ||
| 10.3 Methods for Glycan Labeling and Imaging | 286 | ||
| 10.3.1 Lectins and Antibodies | 287 | ||
| 10.3.2 Metabolic Glycan Labeling | 288 | ||
| 10.3.3 Chemoenzymatic Labeling | 290 | ||
| 10.4 Imaging Protein Glycosylation | 291 | ||
| 10.4.1 General Principles of the Dual-Labeling-Based Methods | 291 | ||
| 10.4.2 FRET-Based Protein-Specific Imaging of Glycosylation | 291 | ||
| 10.4.3 PLA-Based Protein-Specific Imaging of Glycosylation | 293 | ||
| 10.4.4 Protein-Specific Imaging of Glycosylation Based on PEBL | 295 | ||
| 10.4.5 Protein-Specific Imaging of Glycosylation Based on SERS | 296 | ||
| 10.5 Conclusion and Perspective | 296 | ||
| References | 296 | ||
| Chapter 11 - Targeting Glycans of HIV Envelope Glycoproteins for Vaccine Design | 300 | ||
| 11.1 The Human Immunodeficiency Virus | 300 | ||
| 11.1.1 Structure, Genome and Viral Lifecycle | 301 | ||
| 11.1.2 Transmission and Pathogenesis | 303 | ||
| 11.1.3 The Viral Envelope is the Main Target for the Immune System | 304 | ||
| 11.1.4 Immune Response | 305 | ||
| 11.1.5 Current Therapies and Steps towards a Vaccine | 307 | ||
| 11.2 The Viral Envelope Spike | 308 | ||
| 11.2.1 Biosynthesis | 308 | ||
| 11.2.2 Structure and Function of Env | 311 | ||
| 11.2.3 The Glycan Shield | 314 | ||
| 11.2.4 Site-specific N-Linked Glycan Analysis | 317 | ||
| 11.2.5 Glycans in Immune Escape | 320 | ||
| 11.3 A Target for Broadly Neutralizing Antibodies | 321 | ||
| 11.3.1 Sites of Vulnerability | 322 | ||
| 11.3.2 Unusual Features of Broadly Neutralizing Antibodies | 323 | ||
| 11.3.3 Development of Broadly Neutralizing Antibodies | 325 | ||
| 11.3.4 Mechanisms of Neutralization | 326 | ||
| 11.3.5 Application in Therapy and Cure Research | 327 | ||
| 11.4 Strategies for Vaccine Design | 328 | ||
| 11.4.1 Challenges | 328 | ||
| 11.4.2 Rational Immunogen Design to Induce Broadly Neutralizing Antibodies | 329 | ||
| 11.4.3 Chemical Biology of Carbohydrate-Based Immunogens | 332 | ||
| 11.4.4 Vaccination Strategies | 333 | ||
| 11.5 Conclusion and Outlook | 334 | ||
| References | 335 | ||
| Chapter 12 - Design, Synthesis and Evaluation of Mucin Glycopeptide Based Cancer Vaccine | 358 | ||
| 12.1 Introduction | 358 | ||
| 12.2 Mucins as the Target of Anti-Tumor Vaccine | 359 | ||
| 12.2.1 Tumor Antigens | 359 | ||
| 12.2.2 Mucins | 360 | ||
| 12.2.3 MUC1 in Normal Cells | 360 | ||
| 12.2.4 MUC1 in Tumor Cells | 361 | ||
| 12.2.5 Differences in the Glycosylation Types of the MUC1 in Tumor Cells | 362 | ||
| 12.2.6 Immunological Foundation for MUC1 as a Tumor Antigen | 363 | ||
| 12.3 Overview of Fully Synthetic Vaccines | 364 | ||
| 12.3.1 Structures of Fully Synthetic Vaccines | 364 | ||
| 12.3.2 Immune Response to Fully Synthetic Vaccines | 364 | ||
| 12.3.3 Antigen Structures of Fully Synthetic Vaccines | 365 | ||
| 12.3.3.1 Tumor-Associated Carbohydrate Antigens (TACAs) | 365 | ||
| 12.3.3.2 Tumor-Associated MUC1 Peptide Antigens | 365 | ||
| 12.3.3.3 Tumor-Associated MUC1 Glycopeptide Antigens | 367 | ||
| 12.3.4 Immune-Stimulating Structures of Fully Synthetic Vaccines | 367 | ||
| 12.3.4.1 Carrier Proteins | 367 | ||
| 12.3.4.2 Helper T-Cell Epitope Peptides | 368 | ||
| 12.3.4.3 Immunostimulants | 368 | ||
| 12.4 Fully Synthetic Vaccines Based on Mucin Glycopeptides | 368 | ||
| 12.4.1 Carrier Protein Vaccines | 368 | ||
| 12.4.1.1 Carrier Protein Vaccines Based on Bovine Serum Albumin | 368 | ||
| 12.4.1.2 Carrier Protein Vaccines Based on Other Proteins | 372 | ||
| 12.4.2 Two-Component Vaccines | 372 | ||
| 12.4.2.1 Two-Component Vaccines Based on T-Helper Cell Epitope | 373 | ||
| 12.4.2.2 Two-Component Vaccines Based on Immunostimulants | 373 | ||
| 12.4.2.3 Other Strategies for Two-Component Vaccines | 381 | ||
| 12.4.3 Three-Component Vaccines | 381 | ||
| 12.4.3.1 Three-Component Vaccines Based on TLR2 Ligands | 381 | ||
| 12.4.3.2 Three-Component Vaccines Based on TLR9 Ligands | 386 | ||
| 12.4.3.3 Other Strategies for Three-Component Vaccines | 386 | ||
| 12.5 Conclusions | 388 | ||
| Acknowledgements | 389 | ||
| References | 389 | ||
| Chapter 13 - Selective Chemical Glycosylation of Therapeutic Proteins | 394 | ||
| 13.1 Introduction | 394 | ||
| 13.2 Glycosylation for Increasing Pharmacokinetics and Stability | 395 | ||
| 13.3 Glycosylation for Increased Protein Targeting to Disease-Affected Tissues | 403 | ||
| 13.4 Glycosylation for Site-Specific Drug Conjugation | 406 | ||
| 13.5 Conclusion | 410 | ||
| Acknowledgements | 410 | ||
| References | 410 | ||
| Subject Index | 415 |