title: Averaging of viral envelope glycoprotein spikes from electron cryotomography reconstructions using Jsubtomo.
abstract: Enveloped viruses utilize membrane glycoproteins on their surface to mediate entry into host cells. Three-dimensional structural analysis of these glycoprotein 'spikes' is often technically challenging but important for understanding viral pathogenesis and in drug design. Here, a protocol is presented for viral spike structure determination through computational averaging of electron cryo-tomography data. Electron cryo-tomography is a technique in electron microscopy used to derive three-dimensional tomographic volume reconstructions, or tomograms, of pleomorphic biological specimens such as membrane viruses in a near-native, frozen-hydrated state. These tomograms reveal structures of interest in three dimensions, albeit at low resolution. Computational averaging of sub-volumes, or sub-tomograms, is necessary to obtain higher resolution detail of repeating structural motifs, such as viral glycoprotein spikes. A detailed computational approach for aligning and averaging sub-tomograms using the Jsubtomo software package is outlined. This approach enables visualization of the structure of viral glycoprotein spikes to a resolution in the range of 20-40 Å and study of the study of higher order spike-to-spike interactions on the virion membrane. Typical results are presented for Bunyamwera virus, an enveloped virus from the family Bunyaviridae. This family is a structurally diverse group of pathogens posing a threat to human and animal health.
authors.affiliation: Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford; juha@strubi.ox.ac.uk.
authors.forename: Juha T
authors.initials: JT
authors.lastname: Huiskonen
authors.affiliation: Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford.
authors.forename: Marie-Laure
authors.initials: ML
authors.lastname: Parsy
authors.affiliation: Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford.
authors.forename: Sai
authors.initials: S
authors.lastname: Li
authors.affiliation: Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford.
authors.forename: David
authors.initials: D
authors.lastname: Bitto
authors.affiliation: Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford.
authors.forename: Max
authors.initials: M
authors.lastname: Renner
authors.affiliation: Oxford Particle Imaging Centre, Division of Structural Biology, Wellcome Trust Centre for Human Genetics, University of Oxford.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Bowden
mesh_terms: D002042:Bunyamwera virus / Q000737:chemistry / Q000378:metabolism; D055032:Electron Microscope Tomography / Q000379:methods*; D006023:Glycoproteins / Q000032:analysis / Q000378:metabolism; D012984:Software; D014759:Viral Envelope Proteins / Q000032:analysis* / Q000378:metabolism
chemical_list: D006023:Glycoproteins; D014759:Viral Envelope Proteins