2016Mio Review

Citation

Miro, K.; Sato, M. & Sato, C. Senda, T. & Maenaka, K. (Eds.) Advanced methods in Structural Biology. Structural Biology and Electron microscopy. Springer, 2016, 275-292

Abstract

Like X-ray crystallography and NMR, electron microscopy (EM) is now widely applied to determine the structure of proteins and their macromolecular complexes. Single-particle analysis (SPA), which reconstructs the three-dimensional (3D) structure of a protein from its EM images using image processing, has an advantage when the target molecule is difficult to crystallize or only a small amount of protein can be obtained. The technique is based on the theory that two-dimensional EM images of a protein contain sufficient information to reconstruct the original 3D structure. SPA was developed when this theory was applied to ribosomes and the coat protein of icosahedral or helical symmetrical viruses. Because SPA does not require protein crystallization, it is widely applicable to the analysis of solubilized membrane proteins or supermolecular complexes. It allows conformational changes undergone by proteins to be documented. Many other EM-based structural analysis techniques are available in addition to SPA. Electron tomography reconstructs the 3D structure of a protein complex or a cell from a series of images recorded by tilting the specimen in the EM column. Electron crystallography can yield the high-resolution structure of proteins crystallized in two dimensions in a lipid bilayer. Atmospheric scanning electron microscopy directly observes cells in aqueous solution and has realized high-throughput immuno-EM of cells without hydrophobic treatment. It can also visualize protein microcrystals in the crystallization buffer.

Links

http://link.springer.com/protocol/10.1007%2F978-4-431-56030-2_15