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E. Coli Methods and Protocols M E T H O D S I N M O L E C U L a R B I O L O G Y UC Irvine UC Irvine Previously Published Works Title Protein Folding Using a Vortex Fluidic Device. Permalink https://escholarship.org/uc/item/2bb550m5 Journal Methods in molecular biology (Clifton, N.J.), 1586 ISSN 1064-3745 Authors Britton, Joshua Smith, Joshua N Raston, Colin L et al. Publication Date 2017 DOI 10.1007/978-1-4939-6887-9_13 License https://creativecommons.org/licenses/by/4.0/ 4.0 Peer reviewed eScholarship.org Powered by the California Digital Library University of California Methods in Molecular Biology 1586 Nicola A. Burgess-Brown Editor Heterologous Gene Expression in E. coli Methods and Protocols M ETHODS IN M OLECULAR B IOLOGY Series Editor John M. Walker School of Life and Medical Sciences University of Hertfordshire Hatfield, Hertfordshire, AL10 9AB, UK For further volumes: http://www.springer.com/series/7651 Heterologous Gene Expression in E. coli Methods and Protocols Edited by Nicola A. Burgess-Brown SGC, Nuffield Department of Clinical Medicine, University of Oxford, Oxford, UK Editor Nicola A. Burgess-Brown SGC Nuffield Department of Clinical Medicine University of Oxford Oxford, UK ISSN 1064-3745 ISSN 1940-6029 (electronic) Methods in Molecular Biology ISBN 978-1-4939-6885-5 ISBN 978-1-4939-6887-9 (eBook) DOI 10.1007/978-1-4939-6887-9 Library of Congress Control Number: 2017934051 © Springer Science+Business Media LLC 2017 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. 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. The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, express or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Printed on acid-free paper This Humana Press imprint is published by Springer Nature The registered company is Springer Science+Business Media LLC The registered company address is: 233 Spring Street, New York, NY 10013, U.S.A. Preface Heterologous gene expression in E. coli has been one of the most widely used methods for generating recombinant proteins for many scientific analyses and still remains the first choice for most laboratories around the world. The ease of use and low cost of production often lead researchers to initially attempt to express their proteins of interest in E. coli rather than opting for a eukaryotic host. Decades of development have seen the variety of methods for expressing genes in E. coli broaden, with improved media and optimized conditions for growth, a choice of promoter systems to regulate expression, fusion tags to aid solubility and purification, and E. coli host strains to accommodate more challenging or toxic proteins. Having worked in the area of protein production for structural genomics for the past 12 years, and also having a requirement to generate human proteins, I have seen a shift from expression of many genes in E. coli to use of the baculovirus expression system using insect cells and more recently to mammalian cells. This revolution from prokaryotic to eukaryotic expression has been visible throughout the protein production field and is largely due to the requirement to obtain specific proteins linked to disease, for functional assays as well as structures, which may be larger, or require machinery to enable specific post-­ translational modifications. It is perhaps important to note, however, that the structural output from the SGC in Oxford today is still ~80% derived from E. coli. This book is aimed at molecular biologists, biochemists, and structural biologists, both from the beginning of their research careers to those in their prime, to give both an historical and modern overview of the methods available to express their genes of interest in this exceptional organism. The topics are largely grouped under four parts: (I) high- throughput cloning, expression screening, and optimization of expression conditions, (II) protein production and solubility enhancement, (III) case studies to produce challenging proteins and specific protein families, and (IV) applications of E. coli expression. This vol- ume provides scientists with a toolbox for designing constructs, tackling expression and solubility issues, handling membrane proteins and protein complexes, and innovative engineering of E. coli. It will hopefully prove valuable both in small laboratories and in higher throughput facilities. I would like to thank all the authors for their contributions and for making this a global effort. Oxford, UK Nicola A. Burgess-Brown v Contents Preface . v Contributors . xi PART I HiGh-ThROUGhpUT CLONiNG, EXpReSSiON ScReeNiNG, AND OpTiMiZATiON 1 Recombinant Protein Expression in E. coli: A Historical Perspective . 3 Opher Gileadi 2 N- and C-Terminal Truncations to Enhance Protein Solubility and Crystallization: Predicting Protein Domain Boundaries with Bioinformatics Tools . 11 Christopher D.O. Cooper and Brian D. Marsden 3 Harnessing the Profinity eXact™ System for Expression and Purification of Heterologous Proteins in E. coli . 33 Yoav Peleg, Vadivel Prabahar, Dominika Bednarczyk, and Tamar Unger 4 ESPRIT: A Method for Defining Soluble Expression Constructs in Poorly Understood Gene Sequences . 45 Philippe J. Mas and Darren J. Hart 5 Optimizing Expression and Solubility of Proteins in E. coli Using Modified Media and Induction Parameters . 65 Troy Taylor, John-Paul Denson, and Dominic Esposito 6 Optimization of Membrane Protein Production Using Titratable Strains of E. coli . 83 Rosa Morra, Kate Young, David Casas-Mao, Neil Dixon, and Louise E. Bird 7 Optimizing E. coli-Based Membrane Protein Production Using Lemo21(DE3) or pReX and GFP-Fusions . 109 Grietje Kuipers, Markus Peschke, Nurzian Bernsel Ismail, Anna Hjelm, Susan Schlegel, David Vikström, Joen Luirink, and Jan-Willem de Gier 8 High Yield of Recombinant Protein in Shaken E. coli Cultures with Enzymatic Glucose Release Medium EnPresso B . 127 Kaisa Ukkonen, Antje Neubauer, Vinit J. Pereira, and Antti Vasala PART II PROTeiN PURiFicATiON AND SOLUBiLiTY ENhANceMeNT 9 A Generic Protocol for Purifying Disulfide-Bonded Domains and Random Protein Fragments Using Fusion Proteins with SUMO3 and Cleavage by SenP2 Protease . 141 Hüseyin Besir vii viii Contents 10 A Strategy for Production of Correctly Folded Disulfide-Rich Peptides in the Periplasm of E. coli . 155 Natalie J. Saez, Ben Cristofori-Armstrong, Raveendra Anangi, and Glenn F. King 11 Split GFP Complementation as Reporter of Membrane Protein Expression and Stability in E. coli : A Tool to Engineer Stability in a LAT Transporter . 181 Ekaitz Errasti-Murugarren, Arturo Rodríguez-Banqueri, and José Luis Vázquez-Ibar 12 Acting on Folding Effectors to Improve Recombinant Protein Yields and Functional Quality . 197 Ario de Marco 13 Protein Folding Using a Vortex Fluidic Device . 211 Joshua Britton, Joshua N. Smith, Colin L. Raston, and Gregory A. Weiss 14 Removal of Affinity Tags with TEV Protease . 221 Sreejith Raran-Kurussi, Scott Cherry, Di Zhang, and David S. Waugh PART III CASe STUDieS TO PRODUce ChALLeNGiNG PROTeiNS AND SpeciFic PROTeiN FAMiLieS 15 Generation of Recombinant N-Linked Glycoproteins in E. coli . 233 Benjamin Strutton, Stephen R.P. Jaffé, Jagroop Pandhal, and Phillip C. Wright 16 Production of Protein Kinases in E. coli . 251 Charlotte A. Dodson 17 Expression of Prokaryotic Integral Membrane Proteins in E. coli . 265 James D. Love 18 Multiprotein Complex Production in E. coli: The SecYEG-­SecDFYajC-­YidC Holotranslocon . 279 Imre Berger, Quiyang Jiang, Ryan J. Schulze, Ian Collinson, and Christiane Schaffitzel 19 Membrane Protein Production in E. coli Lysates in Presence of Preassembled Nanodiscs . 291 Ralf-Bernhardt Rues, Alexander Gräwe, Erik Henrich, and Frank Bernhard 20 Not Limited to E. coli: Versatile Expression Vectors for Mammalian Protein Expression . 313 Katharina Karste, Maren Bleckmann, and Joop van den Heuvel 21 A Generic Protocol for Intracellular Expression of Recombinant Proteins in Bacillus subtilis . 325 Trang Phan, Phuong Huynh, Tuom Truong, and Hoang Nguyen PART IV AppLicATiONS OF E. COLI EXpReSSiON 22 In Vivo Biotinylation of Antigens in E. coli . 337 Susanne Gräslund, Pavel Savitsky, and Susanne Müller-Knapp Contents ix 23 Cold-Shock Expression System in E. coli for Protein NMR Studies . 345 Toshihiko Sugiki, Toshimichi Fujiwara, and Chojiro Kojima 24 High-Throughput Production of Proteins in E. coli for Structural Studies . 359 Charikleia Black, John J. Barker, Richard B. Hitchman, Hok Sau Kwong, Sam Festenstein, and Thomas B. Acton 25 Mass Spectrometric Analysis of Proteins . 373 Rod Chalk 26 How to Determine Interdependencies of Glucose and Lactose Uptake Rates for Heterologous Protein
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