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Ab initio structure determination from electron microscopic images of single molecules coexisting in different functional states
KTH, School of Technology and Health (STH), Structural Biotechnology.
2010 (English)In: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 18, no 7, 777-786 p.Article in journal (Refereed) Published
Abstract [en]

We have developed methods for ab initio three-dimensional (3D) structure determination from projection images of randomly oriented single molecules coexisting in multiple functional states, to aid the study of complex samples of macromolecules and nanoparticles by electron microscopy (EM). New algorithms for the determination of relative 3D orientations and conformational state assignment of single-molecule projection images are combined with well-established techniques for alignment and statistical image analysis. We describe how the methodology arrives at homogeneous groups of images aligned in 3D and discuss application to experimental EM data sets of the Escherichia coli ribosome and yeast RNA polymerase II.

Place, publisher, year, edition, pages
2010. Vol. 18, no 7, 777-786 p.
Keyword [en]
Proteins, RNA-POLYMERASE-II, CRYO-EM REVEALS, COMMON-LINES, 3-DIMENSIONAL RECONSTRUCTION, CONFORMATIONAL FLEXIBILITY, ORIENTATION REFINEMENT, BIOLOGICAL PARTICLES, 3-D RECONSTRUCTION, PROJECTIONS, RESOLUTION
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:kth:diva-27194DOI: 10.1016/j.str.2010.06.001ISI: 000280046300005PubMedID: 20637414Scopus ID: 2-s2.0-77954629363OAI: oai:DiVA.org:kth-27194DiVA: diva2:375735
Funder
Swedish Research Council
Note
QC 20101209Available from: 2010-12-09 Created: 2010-12-09 Last updated: 2017-12-11Bibliographically approved
In thesis
1. Towards unbiased 3D reconstruction: in single-particle cryo-electron microscopy
Open this publication in new window or tab >>Towards unbiased 3D reconstruction: in single-particle cryo-electron microscopy
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Cryo-electron microscopy of freestanding molecules (single-particles) plays a pivotal role in the difficult and pressing challenge of determining the structures of large macromolecular complexes. Molecular volumes are generated by aligning large sets of randomly oriented two-dimensional (2D) projection images in three dimensions (3D) before reconstruction is performed using tomographic techniques. The increasing popularity of the single-particle method is highly correlated with technical advances in instrumentation and computation. This thesis introduces new computational methods for 3D structure determination from electron microscopic projection images of single molecules. The algorithms have been developed to fill a gap in the single particle methodology – the lack of methods for ab initio 3D reconstruction of asymmetrical or low-symmetry molecules co-existing in different functional states. The proposed approach does not rely on a priori information about the structure or the character of the sample heterogeneity, which minimizes template dependence and makes the methods applicable to a wide range of single molecules. The presented algorithms constitute the basis of a new open source software package - SIMPLE (Single-particle IMage Processing Linux Engine). SIMPLE is an efficient and easy-to-use image processing system for semi-automated ab initio 3D reconstruction from challenging single-particle data sets (asymmetrical particles, significant degree of heterogeneity).

Place, publisher, year, edition, pages
Stockholm: KTH, 2010. 38 p.
Series
Trita-STH : report, ISSN 1653-3836 ; 2010:7
National Category
Industrial Biotechnology
Identifiers
urn:nbn:se:kth:diva-27612 (URN)978-91-7415-837-3 (ISBN)
Public defence
2011-01-14, Föreläsningssalen, Novum, Hälsovägen 7, Huddinge, 10:00 (English)
Opponent
Supervisors
Note
QC 20101214Available from: 2010-12-14 Created: 2010-12-14 Last updated: 2010-12-16Bibliographically approved

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