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Electron Cryomicroscopy and Three-dimensional Computer Reconstruction of Biological Molecules

  1. Steven J Ludtke,
  2. Wah Chiu

Published Online: 15 JUN 2011

DOI: 10.1002/9780470015902.a0002987.pub2

eLS

eLS

How to Cite

Ludtke, S. J. and Chiu, W. 2011. Electron Cryomicroscopy and Three-dimensional Computer Reconstruction of Biological Molecules. eLS. .

Author Information

  1. Baylor College of Medicine, Houston, Texas, USA

Publication History

  1. Published Online: 15 JUN 2011

Abstract

Electron cryomicroscopy (cryo-EM) single-particle analysis is a technique for imaging individual molecules or macromolecules under native-like conditions. When combined with complex computer algorithms, the resulting images can produce a three-dimensional (3D) structure at resolutions now beyond 4 Å. At these resolutions, the protein backbone can be traced, and in many cases sufficient side chains identified to build a full atomistic model akin to that produced by X-ray crystallography. Cryo-EM can also approach problems, which are difficult or impossible for X-ray diffraction or NMR. For example, cryo-EM can be used to characterise the structural flexibility/dynamics of large macromolecules, which is not possible via any other current method. Cryo-EM is also often the method of choice for studying large membrane proteins, which are generally difficult to crystallise. Additional specialised methods exist for 2D crystals or extended structures with helical symmetry.

Key Concepts:

  • Owing to the short wavelength of high voltage electrons, it is possible to resolve features at resolutions ∼1000× higher than light microscopes.

  • Liquid specimens cannot be used under the high vacuum of the microscope and must be either frozen in vitreous ice or chemically fixed before imaging.

  • Tens of thousands of randomly oriented 2D images are combined to produce a high-resolution 3D volumetric structure.

Keywords:

  • electron microscopy;
  • 3D reconstruction;
  • single particle;
  • macromolecule;
  • crystal