title: In Situ Structural Analysis of the Spirochetal Flagellar Motor by Cryo-Electron Tomography.
abstract: The bacterial flagellar motor is a large multi-component molecular machine. Structural determination of such a large complex is often challenging and requires extensive structural analysis in situ. Cryo-electron tomography (cryo-ET) has emerged as a powerful technique that enables us to visualize intact flagellar motors in cells with unprecedented details. Here, we detail the procedure beginning with sample preparation, followed by data acquisition, tomographic reconstruction, sub-tomogram analysis, and ultimately visualization of the intact spirochetal flagellar motor in Borrelia burgdorferi. The procedure is applicable to visualize other molecular machinery in bacteria or other organisms.
authors.affiliation: Department of Pathology and Laboratory Medicine, McGovern Medical School at UTHealth, 6431 Fannin Street, MSB 2.228, Houston, TX, 77030, USA.
authors.forename: Shiwei
authors.initials: S
authors.lastname: Zhu
authors.affiliation: Department of Pathology and Laboratory Medicine, McGovern Medical School at UTHealth, 6431 Fannin Street, MSB 2.228, Houston, TX, 77030, USA.
authors.forename: Zhuan
authors.initials: Z
authors.lastname: Qin
authors.affiliation: Department of Pathology and Laboratory Medicine, McGovern Medical School at UTHealth, 6431 Fannin Street, MSB 2.228, Houston, TX, 77030, USA.
authors.forename: Juyu
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Pathology and Laboratory Medicine, McGovern Medical School at UTHealth, 6431 Fannin Street, MSB 2.228, Houston, TX, 77030, USA.
authors.forename: Dustin R
authors.initials: DR
authors.lastname: Morado
authors.affiliation: Department of Pathology and Laboratory Medicine, McGovern Medical School at UTHealth, 6431 Fannin Street, MSB 2.228, Houston, TX, 77030, USA. jun.liu.1@uth.tmc.edu.
authors.forename: Jun
authors.initials: J
authors.lastname: Liu
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism*; D025065:Borrelia burgdorferi / Q000378:metabolism; D020285:Cryoelectron Microscopy / Q000379:methods; D055032:Electron Microscope Tomography / Q000379:methods; D005407:Flagella / Q000378:metabolism; D020409:Molecular Motor Proteins / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D020409:Molecular Motor Proteins
keywords: Cryo-electron tomography; Flagellar motor; Periplasmic flagellum; Spirochete; Sub-tomogram averaging