Peptides: Chemistry and Biology. Norbert Sewald, Hans-Dieter Jakubke Copyright © 2002 Wiley-VCH Verlag GmbH & Co. KGaA ISBNs: 3-527-30405-3 (Hardback); 3-527-60068-X (Electronic) Norbert Sewald and Hans-Dieter Jakubke Peptides: Chemistry and Biology www.ShimiPedia.ir Peptides: Chemistry and Biology. Norbert Sewald, Hans-Dieter Jakubke Copyright © 2002 Wiley-VCH Verlag GmbH & Co. KGaA ISBNs: 3-527-30405-3 (Hardback); 3-527-60068-X (Electronic) Norbert Sewald and Hans-Dieter Jakubke Peptides: Chemistry and Biology www.ShimiPedia.ir Peptides: Chemistry and Biology. Norbert Sewald, Hans-Dieter Jakubke Copyright © 2002 Wiley-VCH Verlag GmbH & Co. KGaA ISBNs: 3-527-30405-3 (Hardback); 3-527-60068-X (Electronic) Prof. Dr. Norbert Sewald n This book was carefully produced. Nevertheless, Universität Bielefeld authors and publisher do not warrant the infor- Fakultät für Chemie mation contained therein to be free of errors. Organ. und Bioorgan. Chemie Readers are advised to keep in mind that state- PO Box 100131 ments, data, illustrations, procedural details or 33501 Bielefeld, Germany other items may inadvertently be inaccurate. Prof. em. Dr. Hans-Dieter Jakubke The use of general descriptive names, registered Universität Leipzig names, trademarks, etc. in this book does not im- Institut für Biochemie ply, even in the absence of a specific statement, Private address: that such names are exempt from the relevant Albert-Richter-Straße 12 protective laws and regulations and therefore free 01465 Dresden-Langebrück, Germany for general use. Library of Congress Card No.: applied for British Library Cataloguing-in-Publication Data Cover A catalogue record for this book is available from The cover picture shows the TPR1 domain of Hop the British Library. in complex with -Gly-Pro-Thr-Ile-Glu-Glu-Val- Die Deutsche Bibliothek – CIP-Cataloguing-in- Asp-OH (GPTIEEVD). TPR domains participate in Publication Data the ordered assembly of Hsp70-Hsp90 multichape- A catalogue record for this publication is available rone complexes. from Die Deutsche Bibliothek. The TPR1 domain of the adaptor protein Hop specifically recognizes the C-terminal heptapep- © WILEY-VCH Verlag GmbH tide -Pro-Thr-Ile-Glu-Glu-Val-Asp-OH (PTIEEVD) D-69469 Weinheim, 2002 of the chaperone Hsp70 while the TPR2A domain All rights reserved (including those of translation of Hop binds the C-terminal pentapeptide -Met- in other languages). No part of this book may be Glu-Glu-Val-Asp-OH (MEEVD) of the chaperone reproduced in any form – by photoprinting, mi- Hsp90. The EEVD motif is conserved in all solu- crofilm, or any other means – nor transmitted or ble forms of eukaryotic Hsp70 and Hsp90 pro- translated into machine language without written teins. permission from the publishers. Registered Peptide binding is mediated with the EEVD motif. names, trademarks, etc. used in this book, even Both carboxy groups of the C-terminal aspartate when not specifically marked as such, are not to anchor the peptide by electrostatic interactions. be considered unprotected by law. The hydrophobic residues located N-terminally Printed in the Federal Republic of Germany within the peptide are critical for specificity. Printed on acid-free paper [C. Scheufler, A. Brinker, G. Bourenkov, S. Pegora- Typesetting ro, L. Moroder, H. Bartunik, F.U. Hartl, I. Moarefi, K+V Fotosatz GmbH, Beerfelden Printing Structure of TPR domain-peptide complexes: criti- betz-druck gmbH, Darmstadt Bookbinding cal elements in the assembly of the Hsp70-Hsp90 J. Schäffer GmbH & Co.KG, multichaperone machine, Cell 2000, 101, 199; PDB Grünstadt entry 1ELW (http://www.rcsb.org/pdb/)] ISBN 3-527-30405-3 www.ShimiPedia.ir Peptides: Chemistry and Biology. Norbert Sewald, Hans-Dieter Jakubke V Copyright © 2002 Wiley-VCH Verlag GmbH & Co. KGaA ISBNs: 3-527-30405-3 (Hardback); 3-527-60068-X (Electronic) Contents Preface XI Abbreviations XIII 1 Introduction and Background 1 2 Fundamental Chemical and Structural Principles 5 2.1 Definitions and Main Conformational Features of the Peptide Bond 5 2.2 Building Blocks, Classification, and Nomenclature 7 2.3 Analysis of the Covalent Structure of Peptides and Proteins 11 2.3.1 Separation and Purification 12 2.3.1.1 Separation Principles 12 2.3.1.2 Purification Techniques 16 2.3.1.3 Stability Problems 18 2.3.1.4 Evaluation of Homogeneity 19 2.3.2 Primary Structure Determination 20 2.3.2.1 End Group Analysis 21 2.3.2.2 Cleavage of Disulfide Bonds 23 2.3.2.3 Analysis of Amino Acid Composition 24 2.3.2.4 Selective Methods of Cleaving Peptide Bonds 25 2.3.2.5 N-Terminal Sequence Analysis (Edman Degradation) 27 2.3.2.6 C-terminal Sequence Analysis 29 2.3.2.7 Mass Spectrometry 30 2.3.2.8 Peptide Ladder Sequencing 32 2.3.2.9 Assignment of Disulfide Bonds and Peptide Fragment Ordering 33 2.3.2.10 Location of Post-Translational Modifications and Bound Cofactors 35 2.4 Three-Dimensional Structure 36 2.4.1 Secondary Structure 36 2.4.1.1 Helix 37 2.4.1.2 b-Sheet 38 2.4.1.3 Turns 39 2.4.1.4 Amphiphilic Structures 41 2.4.2 Tertiary Structure 43 2.4.2.1 Structure Prediction 46 www.ShimiPedia.ir VI Contents 2.5 Methods of Structural Analysis 47 2.5.1 Circular Dichroism 48 2.5.2 Infrared Spectroscopy 49 2.5.3 NMR Spectroscopy 50 2.5.4 X-Ray Crystallography 52 2.5.5 UV Fluorescence Spectroscopy 54 2.6 References 55 3 Biologically Active Peptides 61 3.1 Occurrence and Biological Roles 61 3.2 Biosynthesis 73 3.2.1 Ribosomal Synthesis 73 3.2.2 Post-translational Modification 76 3.2.2.1 Enzymatic Cleavage of Peptide Bonds 76 3.2.2.2 Hydroxylation 78 3.2.2.3 Carboxylation 78 3.2.2.4 Glycosylation 78 3.2.2.5 Amidation 83 3.2.2.6 Phosphorylation 83 3.2.2.7 Lipidation 85 3.2.2.8 Pyroglutamyl Formation 86 3.2.2.9 Sulfatation 87 3.2.3 Nonribosomal Synthesis 88 3.3 Selected Bioactive Peptide Families 90 3.3.1 Peptide and Protein Hormones 90 3.3.1.1 Liberins and Statins 92 3.3.1.2 Pituitary Hormones 96 3.3.1.3 Neurohypophyseal Hormones 98 3.3.1.4 Gastrointestinal Hormones 99 3.3.1.5 Pancreatic Islet Hormones 100 3.3.1.6 Further Physiologically Relevant Peptide Hormones 103 3.3.2 Neuropeptides 107 3.3.2.1 Opioid Peptides 109 3.3.2.2 Tachykinins 114 3.3.2.3 Further Selected Neuroactive Peptides 116 3.3.3 Peptide Antibiotics 119 3.3.3.1 Nonribosomally Synthesized Peptide Antibiotics 119 3.3.3.2 Ribosomally Synthesized Peptide Antibiotics 124 3.3.4 Peptide Toxins 126 3.4 References 130 4 Peptide Synthesis 135 4.1 Principles and Objectives 135 4.1.1 Main Targets of Peptide Synthesis 135 4.1.1.1 Confirmation of Suggested Primary Structures 135 www.ShimiPedia.ir Contents VII 4.1.1.2 Design of Bioactive Peptide Drugs 136 4.1.1.3 Preparation of Pharmacologically Active Peptides and Proteins 137 4.1.1.4 Synthesis of Model Peptides 138 4.1.2 Basic Principles of Peptide Bond Formation 139 4.2 Protection of Functional Groups 142 a 4.2.1 N -Amino Protection 143 4.2.1.1 Alkoxycarbonyl-Type (Urethane-Type) Protecting Groups 143 4.2.1.2 Carboxamide-Type Protecting Groups 152 4.2.1.3 Sulfonamide and Sulfenamide-Type Protecting Groups 152 4.2.1.4 Alkyl-Type Protecting Groups 153 a 4.2.2 C -Carboxy Protection 154 4.2.2.1 Esters 155 4.2.2.2 Amides and Hydrazides 157 a 4.2.3 C-terminal and Backbone N -Carboxamide Protection 160 4.2.4 Side-chain Protection 162 4.2.4.1 Guanidino Protection 162 4.2.4.2 x-Amino Protection 165 4.2.4.3 x-Carboxy Protection 166 4.2.4.4 Thiol Protection 168 4.2.4.5 Imidazole Protection 171 4.2.4.6 Hydroxy Protection 174 4.2.4.7 Thioether Protection 176 4.2.4.8 Indole Protection 177 4.2.4.9 x-Amide Protection 178 4.2.5 Enzyme-labile Protecting Groups 180 a 4.2.5.1 Enzyme-labile N -Amino Protection 181 a 4.2.5.2 Enzyme-labile C -Carboxy Protection and Enzyme-labile Linker Moieties 182 4.2.6 Protecting Group Compatibility 184 4.3 Peptide Bond Formation 184 4.3.1 Acyl Azides 185 4.3.2 Anhydrides 186 4.3.2.1 Mixed Anhydrides 187 4.3.2.2 Symmetrical Anhydrides 189 4.3.2.3 N-Carboxy Anhydrides 190 4.3.3 Carbodiimides 191 4.3.4 Active Esters 195 4.3.5 Acyl Halides 200 4.3.6 Phosphonium Reagents 201 4.3.7 Uronium Reagents 202 4.3.8 Further Special Methods 204 4.4 Racemization During Synthesis 205 4.4.1 Direct Enolization 205 4.4.2 5(4H)-Oxazolone Mechanism 205 4.4.3 Racemization Tests: Stereochemical Product Analysis 208 www.ShimiPedia.ir VIII Contents 4.5 Solid-Phase Peptide Synthesis (SPPS) 209 4.5.1 Solid Supports and Linker Systems 212 4.5.2 Safety-Catch Linkers 220 4.5.3 Protection Schemes 224 4.5.3.1 Boc/Bzl-protecting Groups Scheme (Merrifield Tactics) 224 4.5.3.2 Fmoc/tBu-protecting Groups Scheme (Sheppard Tactics) 225 4.5.3.3 Three- and More-Dimensional Orthogonality 227 4.5.4 Chain Elongation 227 4.5.4.1 Coupling Methods 227 4.5.4.2 Undesired Problems During Elongation 228 4.5.4.3 Difficult Sequences 230 4.5.4.4 On-Resin Monitoring 232 4.5.5 Automation of the Process 232 4.5.6 Special Methods 233 4.5.7 Peptide Cleavage from the Resin 235 4.5.7.1 Acidolytic Methods 235 4.5.7.2 Side reactions 236 4.5.7.3 Advantages and Disadvantages of the Boc/Bzl and Fmoc/tBu Schemes 237 4.5.8 Examples of Syntheses by Linear SPPS 237 4.6 Biochemical Synthesis 238 4.6.1 Recombinant DNA Techniques 239 4.6.1.1 Principles of DNA Technology 239 4.6.1.2 Examples of Synthesis by Genetic Engineering 243 4.6.1.3 Cell-free Translation Systems 244 4.6.2 Enzymatic Peptide Synthesis 247 4.6.2.1 Introduction 247 4.6.2.2 Approaches to Enzymatic Synthesis 248 4.6.2.3 Manipulations to Suppress Competitive Reactions 250 4.6.2.4 Irreversible C–N Ligations by Mimicking Enzyme Specificity 251 4.6.3 Antibody-catalyzed Peptide Bond Formation 253 4.7 References