Document Detail

Protein secondary structure determination by constrained single-particle cryo-electron tomography.
MedLine Citation:
PMID:  23217682     Owner:  NLM     Status:  MEDLINE    
Cryo-electron microscopy (cryo-EM) is a powerful technique for 3D structure determination of protein complexes by averaging information from individual molecular images. The resolutions that can be achieved with single-particle cryo-EM are frequently limited by inaccuracies in assigning molecular orientations based solely on 2D projection images. Tomographic data collection schemes, however, provide powerful constraints that can be used to more accurately determine molecular orientations necessary for 3D reconstruction. Here, we propose "constrained single-particle tomography" as a general strategy for 3D structure determination in cryo-EM. A key component of our approach is the effective use of images recorded in tilt series to extract high-resolution information and correct for the contrast transfer function. By incorporating geometric constraints into the refinement to improve orientational accuracy of images, we reduce model bias and overrefinement artifacts and demonstrate that protein structures can be determined at resolutions of ∼8 Å starting from low-dose tomographic tilt series.
Alberto Bartesaghi; Federico Lecumberry; Guillermo Sapiro; Sriram Subramaniam
Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, Non-P.H.S.    
Journal Detail:
Title:  Structure (London, England : 1993)     Volume:  20     ISSN:  1878-4186     ISO Abbreviation:  Structure     Publication Date:  2012 Dec 
Date Detail:
Created Date:  2012-12-11     Completed Date:  2013-05-17     Revised Date:  2013-12-11    
Medline Journal Info:
Nlm Unique ID:  101087697     Medline TA:  Structure     Country:  United States    
Other Details:
Languages:  eng     Pagination:  2003-13     Citation Subset:  IM    
Copyright Information:
Copyright © 2012 Elsevier Ltd. All rights reserved.
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MeSH Terms
Chaperonin 60 / chemistry,  ultrastructure
Cryoelectron Microscopy / methods*
Electron Microscope Tomography / methods*
Escherichia coli Proteins / chemistry,  ultrastructure
Models, Molecular*
Protein Structure, Quaternary
Protein Structure, Secondary
Protein Structure, Tertiary
RNA Polymerase II / chemistry,  ultrastructure
Saccharomyces cerevisiae Proteins / chemistry,  ultrastructure
Transcription Factor TFIIB / chemistry,  ultrastructure
Grant Support
Reg. No./Substance:
0/Chaperonin 60; 0/Escherichia coli Proteins; 0/SUA7 protein, S cerevisiae; 0/Saccharomyces cerevisiae Proteins; 0/Transcription Factor TFIIB; EC 2.7.7.-/RNA Polymerase II

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