Near-atomic resolution using electron cryomicroscopy and single-particle reconstruction
- Xing Zhang*,
- Ethan Settembre†,
- Chen Xu‡,
- Philip R. Dormitzer†,§,
- Richard Bellamy¶,
- Stephen C. Harrison†,‖,**, and
- Nikolaus Grigorieff*,**
- *Department of Biochemistry and Howard Hughes Medical Institute, and
- ‡ Rosenstiel Basic Medical Sciences Research Center, Brandeis University, MS029, 415 South Street, Waltham, MA 02454;
- †Laboratory of Molecular Medicine, and
- ‖Howard Hughes Medical Institute, Children's Hospital, 320 Longwood Avenue, Boston, MA 02115; and
- ¶School of Biological Sciences, University of Auckland, Auckland, New Zealand
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Contributed by Stephen C. Harrison, December 10, 2007 (received for review November 26, 2007)
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-Å 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.
Footnotes
- **To whom correspondence may be addressed. E-mail: harrison{at}crystal.harvard.edu or niko{at}brandeis.edu
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Author contributions: P.R.D., S.C.H., and N.G. designed research; X.Z., E.S., and C.X. performed research; E.S., P.R.D., and R.B. contributed new reagents/analytic tools; X.Z., C.X., S.C.H., and N.G. analyzed data; and S.C.H. and N.G. wrote the paper.
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↵ §Present address: Novartis Vaccines and Diagnostics, 350 Massachusetts Avenue, Cambridge, MA 02139.
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Conflict of interest statement: P.R.D. is currently employed by Novartis Vaccines and Diagnostics.
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Data deposition: Density maps of DLP and the nonicosahedrally averaged VP6 structure have been deposited in the Macromolecular Structure Database at the European Bioinformatics Institute, www.ebi.ac.uk/msd-srv/emsearch/index.html (accession codes 1460 and 1461, respectively).
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See Commentary on page 1779.
- © 2008 by The National Academy of Sciences of the USA
