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Proc Natl Acad Sci U S A. 2008 Feb 12;105(6):1867-72. doi: 10.1073/pnas.0711623105. Epub 2008 Jan 31.

Near-atomic resolution using electron cryomicroscopy and single-particle reconstruction.

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  • 1Department of Biochemistry and Howard Hughes Medical Institute, Brandeis University, MS029, 415 South Street, Waltham, MA 02454, USA.

Abstract

Electron cryomicroscopy (cryo-EM) yields images of macromolecular assemblies and their components, from which 3D structures can be determined, by using an image processing method commonly known as "single-particle reconstruction." During the past two decades, this technique has become an important tool for 3D structure determination, but it generally has not been possible to determine atomic models. In principle, individual molecular images contain high-resolution information contaminated by a much higher level of noise. In practice, it has been unclear whether current averaging methods are adequate to extract this information from the background. We present here a reconstruction, obtained by using recently developed image processing methods, of the rotavirus inner capsid particle ("double-layer particle" or DLP) at a resolution suitable for interpretation by an atomic model. The result establishes single-particle reconstruction as a high-resolution technique. We show by direct comparison that the cryo-EM reconstruction of viral protein 6 (VP6) of the rotavirus DLP is similar in clarity to a 3.8-A resolution map obtained from x-ray crystallography. At this resolution, most of the amino acid side chains produce recognizable density. The icosahedral symmetry of the particle was an important factor in achieving this resolution in the cryo-EM analysis, but as the size of recordable datasets increases, single-particle reconstruction also is likely to yield structures at comparable resolution from samples of much lower symmetry. This potential has broad implications for structural cell biology.

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PMID:
18238898
[PubMed - indexed for MEDLINE]
PMCID:
PMC2542862
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