Structural Virology (Rsc Biomolecular Sciences, 21) - Hardcover

Hardcover

ISBN 10: 0854041710 ISBN 13: 9780854041718

Verlag: Royal Society of Chemistry, 2010

Inhaltsangabe

Over the last ten years, much effort has been devoted to improving the biophysical techniques used in the study of viruses. This has resulted in the visualization of these large macromolecular assemblages at atomic level, thus providing the platform for functional interpretation and therapeutic design. Structural Virology covers a wide range of topics and is split into three sections. The first discusses the vast biophysical methodologies used in structural virology, including sample production and purification, confocal microscopy, mass spectrometry, negative-stain and cryo-electron microscopy, X-ray crystallography and nuclear magnetic resonance spectroscopy. The second discusses the role of virus capsid protein structures in determining the functional roles required for receptor recognition, cellular entry, capsid assembly, genome packaging and mechanisms of host immune system evasion. The last section discusses therapeutic strategies based on virus protein structures, including the design of antiviral drugs and the development of viral capsids as vehicles for foreign gene delivery. Each topic covered will begin with a review of the current literature followed by a more detailed discussion of experimental procedures, a step in the viral life cycle, or strategies for therapeutic development. With contributions from experts in the field of structural biology and virology this exceptional monograph will appeal to biomedical scientists involved in basic and /or applied research on viruses. It also provides up-to-date reference material for students entering the field of structural virology as well as scientists already familiar with the area.

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Über die Autorinnen und Autoren

Mavis Agbandje-McKenna is currently Director of the Center for Structural Biology and Associate Professor at the University of Florida College of Medicine. She obtained her BSc (Hons) in Human Biology and Chemistry from the University of Hertfordshire and a PhD in Biophysics at the University of London Institute of Cancer Research. Her postdoctoral research was at Purdue University before she was appointed Senior Research Fellow at the University of Warwick, UK. Robert McKenna obtained his BSc (Physics and Biology) and PhD in Crystallography from the University of London. His postdoctoral research was at Purdue University before he accepted the post of Warwick Research Fellow in the Department of Biological Sciences at the University of Warwick. He is currently Associate Professor at the University of Florida College of Medicine.

Mavis Agbandje-McKenna is currently Director of the Center for Structural Biology and Associate Professor at the University of Florida College of Medicine. She obtained her BSc (Hons) in Human Biology and Chemistry from the University of Hertfordshire and a PhD in Biophysics at the University of London Institute of Cancer Research. Her postdoctoral research was at Purdue University before she was appointed Senior Research Fellow at the University of Warwick, UK. Robert McKenna obtained his BSc (Physics and Biology) and PhD in Crystallography from the University of London. His postdoctoral research was at Purdue University before he accepted the post of Warwick Research Fellow in the Department of Biological Sciences at the University of Warwick. He is currently Associate Professor at the University of Florida College of Medicine.

Von der hinteren Coverseite

Over the last ten years, much effort has been devoted to improving the biophysical techniques used in the study of viruses. This has resulted in the visualization of these large macromolecular assemblages at atomic level, thus providing the platform for functional interpretation and therapeutic design. Structural Virology covers a wide range of topics and is split into three sections. The first discusses the vast biophysical methodologies used in structural virology, including sample production and purification, confocal microscopy, mass spectrometry, negative-stain and cryo-electron microscopy, X-ray crystallography and nuclear magnetic resonance spectroscopy. The second discusses the role of virus capsid protein structures in determining the functional roles required for receptor recognition, cellular entry, capsid assembly, genome packaging and mechanisms of host immune system evasion. The last section discusses therapeutic strategies based on virus protein structures, including the design of antiviral drugs and the development of viral capsids as vehicles for foreign gene delivery. Each topic covered will begin with a review of the current literature followed by a more detailed discussion of experimental procedures, a step in the viral life cycle, or strategies for therapeutic development. With contributions from experts in the field of structural biology and virology this exceptional monograph will appeal to biomedical scientists involved in basic and /or applied research on viruses. It also provides up-to-date reference material for students entering the field of structural virology as well as scientists already familiar with the area.

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Structural Virology

By Mavis Agbandje-McKenna, Robert McKenna

The Royal Society of Chemistry

Copyright © 2011 Royal Society of Chemistry
All rights reserved.

ISBN: 978-0-85404-171-8

Contents

SECTION 1, 
Chapter 1 Production and Purification of Viruses for Structural Studies Brittney L. Gurda and Mavis Agbandje-McKenna, 3, 
Chapter 2 Microscopic Analysis of Viral Cell Binding, Entry and Infection in Live Cells Colin R. Parrish, 22, 
Chapter 3 Probing Viral Capsids in Solution Brian Bothner and Jonathan K. Hilmer, 41, 
Chapter 4 Three-dimensional Structures of Pleiomorphic Viruses from Cryo-Electron Tomography Alasdair C. Steven, Giovanni Cardone, Carmen Butan, Dennis C. Winkler and J. Bernard Heymann, 62, 
Chapter 5 Structure Determination of Icosahedral Viruses Imaged by Cryo-electron Microscopy Robert S. Sinkovits and Timothy S. Baker, 81, 
Chapter 6 X-ray Crystallography of Virus Capsids Lakshmanan Govindasamy, Mavis Agbandje-McKenna and Robert McKenna, 100, 
Chapter 7 Structural Studies of Viral Proteins – X-ray Crystallography John Domsic and Robert McKenna, 123, 
Chapter 8 Solution NMR Spectroscopy in Characterizing Structure, Dynamics and Intermolecular Interactions of Retroviral Structural Proteins Kang Chen and Nico Tjandra, 137, 
SECTION 2, 
Chapter 9 Evolution of Viral Capsid Structures – the Three Domains of Life Reza Khayat and John E. Johnson, 163, 
Chapter 10 Mechanisms of Icosahedral Virus Assembly Adam Zlotnick and Bentley A. Fane, 180, 
Chapter 11 Mechanisms of Genome Packaging Mark Oram and Lindsay W. Black, 203, 
Chapter 12 Attachment and Entry: Receptor Recognition in Viral Pathogenesis Damian C. Ekiert and Ian A. Wilson, 220, 
Chapter 13 Attachment and Entry: Viral Cell Fusion Rachel M. Schowalter, Everett C. Smith and Rebecca Ellis Dutch, 243, 
Chapter 14 Structural Studies on Antibody – Virus Complexes Thomas J. Smith, 261, 
SECTION 3, 
Chapter 15 Development of Anti-HIV Drugs Roxana M. Coman and Robert McKenna, 293, 
Chapter 16 Design of Capsid-binding Antiviral Agents Against Human Rhinoviruses Chuan Xiao, Mark A. McKinlay and Michael G. Rossmann, 319, 
Chapter 17 Viral Vectors for Gene Delivery David J. Dismuke, Steven J. Gray, Matthew L. Hirsch, Richard Samulski and Nicholas Muzyczka, 338, 
Subject Index, 358, 

CHAPTER 1

Production and Purification of Viruses for Structural Studies

BRITTNEY L. GURDA AND MAVIS AGBANDJEMCKENNA

Department of Biochemistry and Molecular Biology, Center for Structural Biology, The McKnight Brain Institute, University of Florida, Gainesville, FL 32610, USA

1 Introduction

Advances in protein production and purification techniques over the past two decades have allowed the structural study of numerous proteins and macromolecular assemblages that would have otherwise been intractable to the necessary approaches (detailed in the following chapters). This chapter focuses on the production and purification of intact viral capsids (particles) with/ without genome for structure determination. The production and purification of viral proteins for structure determination by X-ray crystallography and NMR spectroscopy are the subjects of Chapters 7 and 8, respectively. Crystallization is often considered a method of purification and a function of purity, often of a protein or virus capsid, and, as such, sample preparation for structure determination by X-ray crystallography places high demands on sample quality. Screening trials to identify the optimal crystallization conditions also require large quantities of sample compared with the majority of other structure determination approaches discussed in the subsequent chapters of this monograph. Virus samples produced for such analyses also have to be both stable and soluble in their storage buffer since degradation and aggregation are detrimental to the crystallization process. Hence this chapter will focus on methodologies to produce and purify virus capsids (Figure 1) in quantities suitable for structure determination by X-ray crystallography, with the premise that such a sample would also be suitable for structural or biophysical analysis using other methodologies.

2 Expression Systems

Most viruses are considered hazardous material in their wild-type (wt) infectious form (for information on safe handling and containment of infectious microorganisms and hazardous biological materials, see http://www.cdc.gov/ biosafety) and are therefore often studied in a recombinant form. Significant effort has been extended into the development of heterologous expression systems to produce recombinant viral proteins which will assemble into viral capsids. The system selected for use is often dependent on the properties of the viral genes and the environmental requirements of the final product. However, the most important factor to consider is the capacity of the host cells to translate the RNA transcript, to ensure proper folding of the gene product and to sustain the protein(s) expressed in an intact and functional state. Protein expression systems contain at least four general components: (1) the genetic elements necessary for transcription/translation and selection; (2) in vector- based systems, a suitable replicon: plasmid, virus genes, etc.; (3) a host strain containing the appropriate genetic traits needed to function with the specific expression signals and selection scheme; and (4) the culturing conditions for the transformed cells or organisms.

Eukaryotic Systems

Mammalian Cells

Since most viruses currently studied are of human or animal origin, mammalian tissue culture is an ideal source to generate viral capsids for structural studies which are generally aimed at functional annotation. In this system, proper folding is achieved and modifications such as complex glycosylation, phosphorylation, acylation, acetylation and γ-carboxylation are obtained. However, yields can be low, depending on gene product(s), ranging from 0.1 to 100 mg L-1 of culture volume. For some of the structural approaches discussed in Section 1 of this monograph, low yields may not be a problem since small amounts of sample are adequate. However, low yields can become problematic in crystallization, especially with a virus that does not have an established crystallization condition. In such a situation, numerous preparation steps may be required to obtain the quantities needed to screen crystallization conditions efficiently. Supplies and reagents can then become expensive, depending on individual cell line requirements. In addition, considerable time and resources can be spent on the construction of a suitable expression system and equally on optimization for suitable yields. In such situations, it is always advisable to seek the expertise of an established molecular biologist before designing new constructs.

Established cell lines and protocols exists for many different tissue systems and, although most of these cell lines are derived from human or mouse tissues, other mammalian cell culture lines are available, such as monkey, raccoon, horse, pig and rabbit. The American Type Culture Collection (ATCC) has over 3400 cell lines from 80 different species, including over 950 cancer cell lines (http://www.atcc.org/). Other cell suppliers include the Health Protection Agency Culture Collections (HPACC; http://www.hpacultures.org.uk/), the German Research Center for Biological Material (DSMZ; http://www.dsmz.de/) and the Riken BioResource Center Cell Bank (Riken;...

Details zur Ausgabe

• Verlag: Royal Society of Chemistry
• Erscheinungsdatum: 2010
• Sprache: Englisch
• ISBN 10: 0854041710
• ISBN 13: 9780854041718
• Einband: Gebundene Ausgabe
• Anzahl der Seiten: 371
• Herausgeber: Agbandje-mckenna Mavis, McKenna Robert
• Kontakt zum Hersteller: Nicht verfügbar
• Verantwortliche Person: Nicht verfügbar