title: Capture and imaging of a prehairpin fusion intermediate of the paramyxovirus PIV5.
abstract: During cell entry, enveloped viruses fuse their viral membrane with a cellular membrane in a process driven by energetically favorable, large-scale conformational rearrangements of their fusion proteins. Structures of the pre- and postfusion states of the fusion proteins including paramyxovirus PIV5 F and influenza virus hemagglutinin suggest that this occurs via two intermediates. Following formation of an initial complex, the proteins structurally elongate, driving a hydrophobic N-terminal "fusion peptide" away from the protein surface into the target membrane. Paradoxically, this first conformation change moves the viral and cellular bilayers further apart. Next, the fusion proteins form a hairpin that drives the two membranes into close opposition. While the pre- and postfusion hairpin forms have been characterized crystallographically, the transiently extended prehairpin intermediate has not been visualized. To provide evidence for this extended intermediate we measured the interbilayer spacing of a paramyxovirus trapped in the process of fusing with solid-supported bilayers. A gold-labeled peptide that binds the prehairpin intermediate was used to stabilize and specifically image F-proteins in the prehairpin intermediate. The interbilayer spacing is precisely that predicted from a computational model of the prehairpin, providing strong evidence for its structure and functional role. Moreover, the F-proteins in the prehairpin conformation preferentially localize to a patch between the target and viral membranes, consistent with the fact that the formation of the prehairpin is triggered by local contacts between F- and neighboring viral receptor-binding proteins (HN) only when HN binds lipids in its target membrane.
authors.affiliation: Department of Chemistry, University of Pennsylvania, Philadelphia, PA 19104, USA.
authors.forename: Yong Ho
authors.initials: YH
authors.lastname: Kim
authors.forename: Jason E
authors.initials: JE
authors.lastname: Donald
authors.forename: Gevorg
authors.initials: G
authors.lastname: Grigoryan
authors.forename: George P
authors.initials: GP
authors.lastname: Leser
authors.forename: Alexander Y
authors.initials: AY
authors.lastname: Fadeev
authors.forename: Robert A
authors.initials: RA
authors.lastname: Lamb
authors.forename: William F
authors.initials: WF
authors.lastname: DeGrado
mesh_terms: D002462:Cell Membrane / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D007150:Immunohistochemistry; D046529:Microscopy, Electron, Transmission; D008954:Models, Biological; D010252:Paramyxoviridae / Q000378:metabolism*; D011487:Protein Conformation; D017510:Protein Folding; D014461:Ultracentrifugation; D014760:Viral Fusion Proteins / Q000378:metabolism*; D053585:Virus Attachment
chemical_list: D014760:Viral Fusion Proteins