A new cryo-EM single-particle Ab initio reconstruction method visualizes secondary structure elements in an ATP-Fueled AAA+ motor

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Abstract

The generation of ab initio three-dimensional (3D) models is a bottleneck in the studies of large macromolecular assemblies by single-particle cryo-electron microscopy. We describe here a novel method, in which established methods for two-dimensional image processing are combined with newly developed programs for joint rotational 3D alignment of a large number of class averages (RAD) and calculation of 3D volumes from aligned projections (VolRec). We demonstrate the power of the method by reconstructing an 660-kDa ATP-fueled AAA+ motor to 7.5 Å resolution, with secondary structure elements identified throughout the structure. We propose the method as a generally applicable automated strategy to obtain 3D reconstructions from unstained single particles imaged in vitreous ice.

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Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Biological Sciences

Keywords

  • electron microscopy, cryo-EM, single particle, reconstruction, alignment
Original languageEnglish
Pages (from-to)934-947
JournalJournal of Molecular Biology
Volume375
Issue number4
Publication statusPublished - 2008
Publication categoryResearch
Peer-reviewedYes