Averaging of viral envelope glycoprotein spikes from electron cryotomography reconstructions using Jsubtomo.

Journal article

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: Huiskonen, J; Parsy, M; Li, S; Bitto, D; Renner, M

• Publisher: Journal of Visualized Experiments
• Journal: Journal of visualized experiments : JoVE
• Issue: 92
• Pages: e51714
• Publication date: 2014-01-01
• DOI: 10.3791/51714
• EISSN: 1940-087X
• ISSN: 1940-087X
• Language: English
• Keywords: Immunology; Glycoprotein spike; Structure; Electron cryo-microscopy; Subtomogram; Membrane virus; Cryo-electron microscopy; Cryo-electron tomography; Issue 92; Averaging; Electron cryo-tomography; Enveloped virus