title: Statistical modeling and removal of lipid membrane projections for cryo-EM structure determination of reconstituted membrane proteins.
abstract: This paper describes steps in the single-particle cryo-EM 3D structure determination of membrane proteins in their membrane environment. Using images of the Kv1.2 potassium-channel complex reconstituted into lipid vesicles, we describe procedures for the merging of focal-pairs of exposures and the removal of the vesicle-membrane signal from the micrographs. These steps allow 3D reconstruction to be performed from the protein particle images. We construct a 2D statistical model of the vesicle structure based on higher-order singular value decomposition (HOSVD), by taking into account the structural symmetries of the vesicles in polar coordinates. Non-roundness in the vesicle structure is handled with a non-linear shape alignment to a reference, which ensures a compact model representation. The results show that the learned model is an accurate representation of the imaged vesicle structures. Precise removal of the strong membrane signals allows better alignment and classification of images of small membrane-protein particles, and allows higher-resolution 3D reconstruction.
authors.affiliation: Department of Computer Science, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen, Denmark.
authors.forename: Katrine Hommelhoff
authors.initials: KH
authors.lastname: Jensen
authors.affiliation: Department of Computer Science, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen, Denmark.
authors.forename: Sami Sebastian
authors.initials: SS
authors.lastname: Brandt
authors.affiliation: Department of Cellular and Molecular Physiology, Yale University School of Medicine, 333 Cedar St., New Haven, CT 06520, USA; RIKEN Center for Life Science Technologies, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa 230-0045, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Shigematsu
authors.affiliation: Department of Cellular and Molecular Physiology, Yale University School of Medicine, 333 Cedar St., New Haven, CT 06520, USA.
authors.forename: Fred J
authors.initials: FJ
authors.lastname: Sigworth
mesh_terms: D000465:Algorithms; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D020285:Cryoelectron Microscopy / Q000379:methods*; D021621:Imaging, Three-Dimensional / Q000379:methods; D051663:Kv1.2 Potassium Channel / Q000378:metabolism / Q000648:ultrastructure*; D008051:Lipid Bilayers / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism / Q000648:ultrastructure*; D015233:Models, Statistical; D025341:Principal Component Analysis; D051381:Rats; D053835:Unilamellar Liposomes / Q000378:metabolism
chemical_list: D051663:Kv1.2 Potassium Channel; D008051:Lipid Bilayers; D008565:Membrane Proteins; D053835:Unilamellar Liposomes
keywords: Higher-order singular value decomposition; Kv1.2; Liposome; Potassium channel; Single-particle reconstruction; Statistical shape modeling
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