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A Practical Guide to Membrane Protein Purification is written especially for researchers who have some familarity with separation of water-soluble proteins, but who may not be aware of the pitfalls they face with membrane proteins. This guide presents techniques in a concise form, emphasizing the aspects unique to membrane proteins. The book explains the principles of the methods, permitting researchers and students new to this area to adapt these techniques to their particular needs. The second volume in the series, this book is an essential manual for investigations of structure and function of native membrane proteins, as well as for purification of these proteins for immunization and protein sequencing. Separation, Detection, and Characterization of Biological Macromolecules is a new series of laboratory guides. Each volume focuses on a topic of central interest to scientists and students in biomedical and biological research. Introductory chapters are followed by clear, step-by-step protocols that present principles and practice. These concise manuals are designed for optimal understanding of methods as well as for practical benchtop use. - Provides general guidelines and strategies for isolation of membrane proteins - Describes detailed practical procedures that have been the widest applications, and lowest specialized equipment needs - Gives special emphasis to new native and denaturing electrophoresis techniques - Explains modifications of techniques used for water-soluble proteins
This second edition of Membrane Protein Purification and Crystallization, A Practical Guide is written for bench scientists working in the fields of biochemistry, biology, and proteomic research. This guide presents isolation and crystallization techniques in a concise form, emphasizing the critical aspects unique to membrane proteins. It explains the principles of the methods and provides protocols of general use, permitting researchers and students new to this area to adapt these techniques to their particular needs. This edition is not only an update but is comprised mainly of new contributions. It is the first monograph compiling the essential approaches for membrane protein crystallization, and emphasizes recent progress in production and purification of recombinant membrane proteins. - Provides general guidelines and strategies for isolation and crystallization of membrane proteins - Gives detailed protocols that have wide application, and low specialized equipment needs - Emphasizes recent progress in production and purification of recombinant membrane proteins, especially of histidine-tagged and other affinity-epitope-tagged proteins - Summarizes recent developments of Blue-Native PAGE, a high resolution separation technique, which is independent of the use of recombinant techniques, and is especially suited for proteomic analyses of membrane protein complexes - Gives detailed protocols for membrane protein crystallization, and describes the production and use of antibody fragments for high resolution crystallization - Presents a comprehensive guide to 2D-crystallization of membrane proteins
Guide to Protein Purification, Second Edition provides a complete update to existing methods in the field, reflecting the enormous advances made in the last two decades. In particular, proteomics, mass spectrometry, and DNA technology have revolutionized the field since the first edition's publication but through all of the advancements, the purification of proteins is still an indispensable first step in understanding their function. This volume examines the most reliable, robust methods for researchers in biochemistry, molecular and cell biology, genetics, pharmacology and biotechnology and sets a standard for best practices in the field. It relates how these traditional and new cutting-edge methods connect to the explosive advancements in the field. This "Guide to" gives imminently practical advice to avoid costly mistakes in choosing a method and brings in perspective from the premier researchers while presents a comprehensive overview of the field today. - Gathers top global authors from industry, medicine, and research fields across a wide variety of disciplines, including biochemistry, genetics, oncology, pharmacology, dermatology and immunology - Assembles chapters on both common and less common relevant techniques - Provides robust methods as well as an analysis of the advancements in the field that, for an individual investigator, can be a demanding and time-consuming process
Designed as a research-level guide to current strategies and methods of membrane protein production on the small to intermediate scale, this practice-oriented book provides detailed, step-by-step laboratory protocols as well as an explanation of the principles behind each method, together with a discussion of its relative advantages and disadvantages. Following an introductory section on current challenges in membrane protein production, the book goes on to look at expression systems, emerging methods and approaches, and protein specific considerations. Case studies illustrate how to select or sample the optimal production system for any desired membrane protein, saving both time and money on the laboratory as well as the technical production scale. Unique in its coverage of "difficult" proteins with large membrane-embedded domains, proteins from extremophiles, peripheral membrane proteins, and protein fragments.
This book examines detailed experimental and computational approaches for the analysis of many aspects vital to the understanding of membrane protein structure and function. Readers will receive guidance on the selection and use of methods for over-expression and purification, tools to characterize membrane proteins within different phospholipid bilayers, direction on functional studies, and approaches to determine the structures of membrane proteins. Detailed experimental steps for specific membrane proteins with critical notes allow the protocols to be modified to different systems. Written for the highly successful Methods in Molecular Biology series, chapters include the kind of practical information and implementation advice that leads to excellent, reproducible results. Authoritative and up-to-date, Structure and Function Studies of Membrane Proteins serves as an ideal guide for biologists, biochemists, and biophysicists striving to further understand these essential proteins and their many biological roles.
This manual contains necessary and useful information and data in an easily accessible format relating to the use of membranes. Membranes are among the most important engineering components in use today, and each year more and more effective uses for membrane technologies are found - for example: water purification, industrial effluent treatment, solvent dehydration by per-vaporation, recovery of volatile organic compounds, protein recovery, bioseparations and many others.The pace of change in the membrane industry has been accelerating rapidly in recent years, occasioned in part by the demand of end-users, but also as a result of the investment in R&D by manufacturers. To reflect these changes the author has obtained the latest information from some of the leading suppliers in the business. In one complete volume this unique handbook gives practical guidance to using selected membrane processes in individual industries while also providing a useful guide to equipment selection and usage.
Why a Second Edition?The Second Edition provides practical answers to the general question, "How can I obtain useful sequence information from my protein or peptide?" rather than the more specific question asked in the first edition, "How can I obtain the N-terminal sequence?" Important new methods include ways of dealing with blocked N termini, computer analysis of protein sequences, and the recent revolution in mass spectrometry. - Mass spectrophotometric characterization of proteins and peptides - N-terminal sequencing of proteins with blocked N termini - Internal amino acid sequence analysis after protease digestion in-gel and on-blot - Improved microscale peptide purification methods - Computer analysis of protein sequences - New protocols tested and refined through everyday use in authors' laboratories - Updated reference chapter covering all aspects of protein microsequencing
It is a truism of science that the more fundamental the subject, the more universally applicable it is. Neverthelens, it is important to strike a level of "fundamentalness" appropriate to the task in hand. For example, an in-depth study of the mechanics of motor cars would tell one nothing about the dynamics of traffic. Traffic exists on a different "level" - it is dependent upon the existence of motor vehicles but the physics and mathematics of traffic can be adequately addressed by considering motor vehicles as mobile "blobs", with no consideration of how they become mobile. To start a discourse on traffic with a consideration of the mechanics of motor vehicles would thus be inappropropriate. In writing this volume, I have wrestled with the question of the appropriate level at which to address the physics underlying many of the techniques used in protein isolation. I have tried to strike a level as would be used by a mechanic (with perhaps a slight leaning towards an engineer) - i.e. a practical level, offering appropriate insight but with minimal mathematics. Some people involved in biochemical research have a minimal grounding in chemistry and physics and so I have tried to keep it as simple as possible.
The precise knowledge of the structure of biological macromolecules forms the basis of understanding their function and their mechanism of action. It also lays the foundation for rational protein and drug design. The only method to obtain this knowledge is still crystallography. At present, the structures of about 400 proteins are known at or nearly at atomic proteins. However, only two of them are membrane proteins or complexes of the membrane proteins. The reasons for the difference is not the crystals of membrane proteins resists forming special problems when being analysed. The reason is that the membrane proteins resist into forming into well-ordered crystals. The intention of this book is to help to produce well-ordered crystals proteins and to provide guidelines, it is aimed at both biochemists and protein crystallographer‘s.