Cryogenic Electron Microscopy and Single-Particle Analysis

About 20 years ago, the first three-dimensional (3D) reconstructions at subnanometer (<10-Å) resolution of an icosahedral virus assembly were obtained by cryogenic electron microscopy (cryo-EM) and single-particle analysis. Since then, thousands of structures have been determined to resolutions ranging from 30 Å to near atomic (<4 Å). Almost overnight, the recent development of direct electron detectors and the attendant improvement in analysis software have advanced the technology considerably. Near-atomicresolution reconstructions can now be obtained, not only for megadalton macromolecular complexes or highly symmetrical assemblies but also for proteins of only a few hundred kilodaltons. We discuss the developments that led to this breakthrough in high-resolution structure determination by cryo-EM and point to challenges that lie ahead. 499 A nn u. R ev . B io ch em . 2 01 5. 84 :4 99 -5 17 . D ow nl oa de d fr om w w w .a nn ua lr ev ie w s. or g A cc es s pr ov id ed b y B ir kb ec k U ni ve rs ity o f L on do n on 0 3/ 25 /1 6. F or p er so na l u se o nl y. BI84CH18-Elmlund ARI 14 April 2015 12:9 Alignment: the process of bringing images into register or of determining their relative 3D orientations Three-dimensional (3D) reconstruction: a 3D Coulomb potential map of the imaged structure, or the process of generating a 3D Coulomb potential map from images Projection image: represents the entire internal structure of the projected object; a medical X-ray image is a common example Maximum likelihood (ML): a method for estimating the parameters of a statistical model