CHAPTER5StructureDeterminationofIcosahedralViruses.pdfVIP

CHAPTER 5 Structure Determination of Icosahedral Viruses Imaged by Cryo-electron Microscopy ROBERT S. SINKOVITSa,c AND TIMOTHY S. BAKERa,b a Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA 92093, USA; b Division of Biological Sciences, University of California, San Diego, La Jolla, CA 92093, USA; c San Diego Supercomputer Center, University of California, San Diego, La Jolla, CA 92093, USA 1 Introduction Three-dimensional (3D) image reconstruction of icosahedral viruses by trans- mission electron microscopy (TEM) began with the pioneering work on negatively stained samples in Cambridge, UK.1 This ushered in a new era for virus structure determination and helped lay a firm foundation for subsequent, near atomic resolution X-ray crystallographic studies of viruses such as tomato bushy stunt2 and southern bean mosaic virus.3 Crowther et al.’s elegant com- mon lines formulation4 was extremely laborious by today’s standards as it required hands-on inspection and analysis of images and their Fourier trans- forms to identify the orientations of individual virus particles on the TEM ˚ support grid. At that time, the B30 A resolution that could be achieved was limited not by the ability to collect and analyze adequate numbers of images but rather by effects of stain, radiation damage and various distortions to the sample. The advent of single-particle cryo-electron microscopy (cryo-EM) subse- quently revolutionized the use of microscopy for structure determination for RSC Biomolecular Sciences No. 21 Structural Virology Edited by Mavis Agbandje-McKenna and Robert McKenna r Royal Society of Chemistry 2011 Published by the Royal Society of Chemistry, 81 82 Chapter 5