M ETHODS IN MOLECULAR BIOLOGY

Transcription

1 M ETHODS IN MOLECULAR BIOLOGY Series Editor John M. Walker School of Life Sciences University of Hertfordshire Hatfield, Hertfordshire, AL10 9AB, UK For further volumes:

2

3 Peptide Synthesis and Applications Second Edition Edited by Knud J. Jensen Department of Chemistry, Faculty of Sciences, University of Copenhagen, Frederiksberg, Denmark Pernille Tofteng Shelton IGM, Faculty of Life Sciences, University of Copenhagen, Zealand Pharma, Glostrup, Denmark Søren L. Pedersen IGM, Faculty of Life Sciences, University of Copenhagen, Gubra, Hørsholm, Denmark

4 Editors Knud J. Jensen Department of Chemistry Faculty of Sciences University of Copenhagen Frederiksberg, Denmark Pernille Tofteng Shelton IGM, Faculty of Life Sciences University of Copenhagen Zealand Pharma, Glostrup, Denmark Søren L. Pedersen IGM, Faculty of Life Sciences University of Copenhagen Gubra, Hørsholm, Denmark ISSN ISSN (electronic) ISBN ISBN (ebook) DOI / Springer New York Heidelberg Dordrecht London Library of Congress Control Number: Springer Science+Business Media New York 2013 This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. Exempted from this legal reservation are brief excerpts in connection with reviews or scholarly analysis or material supplied specifically for the purpose of being entered and executed on a computer system, for exclusive use by the purchaser of the work. Duplication of this publication or parts thereof is permitted only under the provisions of the Copyright Law of the Publisher s location, in its current version, and permission for use must always be obtained from Springer. Permissions for use may be obtained through RightsLink at the Copyright Clearance Center. Violations are liable to prosecution under the respective Copyright Law. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. While the advice and information in this book are believed to be true and accurate at the date of publication, neither the authors nor the editors nor the publisher can accept any legal responsibility for any errors or omissions that may be made. The publisher makes no warranty, express or implied, with respect to the material contained herein. Printed on acid-free paper Humana Press is a brand of Springer Springer is part of Springer Science+Business Media (

5 Pref ace Protocols for Solid-Phase Peptide Synthesis Peptides are used ubiquitously for studies in biology, biochemistry, chemical biology, peptide- based medicinal chemistry, and indeed many other areas of research. There is a solid number of marketed peptide drugs, and the prospects for the development of new peptide drugs are very encouraging. Most of these peptides are prepared by chemical synthesis, where solid-phase peptide synthesis is the predominant method for preparation of peptides on a laboratory scale and increasingly also on an industrial scale. Practical methodologies for peptide synthesis are the focus of this book. Thus not all reported methods could be described in length. The aim of this book is to provide laboratory protocols for both the specialist and the nonspecialist. The basic protocols provided here for solid-phase peptide synthesis are intended as a practical introduction to peptide synthesis, while the chapters on posttranslational and other modifications hopefully will also appeal to experienced peptide chemists. The first chapter provides an introduction to the basic concepts in peptide synthesis and provides a starting (reference) point for the nonspecialist. The subsequent chapters provide protocols based on experience from the contributors laboratories. It commences with basic protocols for the synthesis of linear, unmodified peptides. The following chapters describe protocols for the synthesis of C-terminally modified peptides, particularly peptide thioesters and aldehydes. This is combined with methods for native chemical ligation and expressed protein ligation. Next, chapters on cyclic peptides and posttranslationally modified peptides describe their synthesis, including in phospho-, glyco-, and lipopeptides. Methods for the assembly of peptidomimetic peptoids are described. Technology is also an important aspect of this book, hence an overview of instruments is provided, which is followed by methods for the relatively new area of microwave heating in peptide synthesis. Finally, the assembly of peptide and glycopeptide microarrays is included in the chapters on glycopeptide synthesis. We have aimed at presenting a broad range of synthetic methods and different approaches to the synthesis of peptides. To achieve this we asked peptide scientists from around the world to contribute protocols based on the chemistries they use in their own laboratories. We thank all the authors for contributing excellent chapters to this book. Frederiksberg, Denmark Zealand Pharma, Glostrup, Denmark Gubra, Hørsholm, Denmark Knud J. Jensen Pernille Tofteng Shelton Søren L. Pedersen v

6

7 Contents Preface Contributors v ix 1 Solid-Phase Peptide Synthesis: An Introduction Knud J. Jensen 2 Linkers, Resins, and General Procedures for Solid-Phase Peptide Synthesis Pernille Tofteng Shelton and Knud J. Jensen 3 Peptide Release, Side-Chain Deprotection, Work-Up, and Isolation Søren L. Pedersen and Knud J. Jensen 4 Synthesis of Peptides Using Tert-Butyloxycarbonyl (Boc) as the α-amino Protection Group Søren W. Pedersen, Christopher J. Armishaw, and Kristian Strømgaard 5 Sequential Formation of Regioselective Disulfide Bonds in Synthetic Peptides with Multiple Disulfide Bonds Fazel Shabanpoor, Mohammed Akhter Hossain, Feng Lin, and John D. Wade 6 Synthesis of Cyclic Disulfide-Rich Peptides Muharrem Akcan and David J. Craik 7 Preparation of C-terminally Modified Chemokines by Expressed Protein Ligation Lars Baumann, Max Steinhagen, and Annette G. Beck-Sickinger 8 Synthesis of C-Terminal Peptide Thioesters Using Fmoc- Based Solid-Phase Peptide Chemistry Pernille Tofteng Shelton and Knud J. Jensen 9 Backbone Amide Linker Strategy: Protocols for the Synthesis of C-Terminal Peptide Aldehydes Pernille Tofteng Shelton and Knud J. Jensen 10 Synthesis of N-Methylated Peptides: On-Resin Methylation and Microwave-Assisted Couplings Renée Roodbeen and Knud J. Jensen 11 Synthesis of Antimicrobial Peptoids Paul R. Hansen and Jens K. Munk 12 Synthesis of Lipidated Peptides Federica Rosi and Gemma Triola 13 Solid-Phase Synthesis of Phosphopeptides Kim B. Højlys-Larsen and Knud J. Jensen vii

8 viii Contents 14 Synthesis of O-Glycopeptides and Construction of Glycopeptide Microarrays Ola Blixt and Emiliano Cló 15 Instruments for Automated Peptide Synthesis Søren L. Pedersen and Knud J. Jensen 16 Microwave-Assisted Solid-Phase Peptide Synthesis Using the Biotage Syro Wave Leila Malik and Knud J. Jensen 17 Microwave-Assisted Solid-Phase Peptide Synthesis Based on the Fmoc Protecting Group Strategy (CEM) Grace S. Vanier Index

9 Contributors MUHARREM AKCAN Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, Australia CHRISTOPHER J. ARMISHAW Torrey Pines Institute for Molecular Studies, Port St. Lucie, FL, USA LARS BAUMANN Institute of Biochemistry, Universität Leipzig, Leipzig, Germany ANNETTE G. BECK-SICKINGER Institute of Biochemistry, Universität Leipzig, Leipzig, Germany OLA BLIXT Department of Chemistry, Faculty of Sciences, University of Copenhagen, Copenhagen, Denmark EMILIANO CLÓ Novo Nordisk A/S, Måløv, Denmark DAVID J. CRAIK Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, Australia PAUL R. HANSEN Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark KIM B. HØJLYS-LARSEN Symphogen A/S, Lyngby, Denmark MOHAMMED AKHTER HOSSAIN Florey Neuroscience Institutes, School of Chemistry, The University of Melbourne, Parkville, VIC, Australia KNUD J. JENSEN Department of Chemistry, Faculty of Sciences, University of Copenhagen, Frederiksberg, Denmark FENG LIN Florey Neuroscience Institutes, The University of Melbourne, Parkville, VIC, Australia LEILA MALIK Department of Chemistry, Faculty of Sciences, University of Copenhagen, Frederiksberg, Denmark JENS K. MUNK Department of Drug Design and Pharmacology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark SØREN L. PEDERSEN IGM, Faculty of Life Sciences, University of Copenhagen, Gubra, Hørsholm, Denmark SØREN W. PEDERSEN Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark RENÉE ROODBEEN Department of Chemistry, Faculty of Sciences, University of Copenhagen, Frederiksberg, Denmark FEDERICA ROSI Abt. Chemische Biologie, Max-Planck-Institut für Molekulare Physiologie, Dortmund, Germany FAZEL SHABANPOOR Florey Neuroscience Institutes, School of Chemistry, The University of Melbourne, Parkville, VIC, Australia MAX STEINHAGEN Institute of Biochemistry, Universität Leipzig, Leipzig, Germany KRISTIAN STRØMGAARD Department of Drug Design and Pharmacology, University of Copenhagen, Copenhagen, Denmark PERNILLE TOFTENG SHELTON IGM, Faculty of Life Sciences, University of Copenhagen, Zealand Pharma, Glostrup, Denmark ix

10 x Contributors GEMMA TRIOLA Abt. Chemische Biologie, Max-Planck-Institut für Molekulare Physiologie, Dortmund, Germany GRACE S. VANIER CEM Corporation, Matthews, NC, USA JOHN D. WADE Florey Neuroscience Institutes, School of Chemistry, The University of Melbourne, Parkville, VIC, Australia