Refolding of a paramyxovirus F protein from prefusion to postfusion conformations observed by liposome binding and electron microscopy.
For paramyxoviruses, two viral glycoproteins are key to the entry process: an attachment protein (HN, H, or G) and the fusion protein (F). The F protein folds to a metastable state that can be triggered to undergo large conformational rearrangements to a fusogenic intermediate and a more stable postfusion state. The triggering mechanism that controls paramyxovirus fusion has not been elucidated. To correlate the molecular structure of a soluble form of the prefusion F (PIV5 F-GCNt) with the biological function of F, soluble F protein was triggered to refold. In the absence of HN, heat was found to function as a surrogate F trigger, and F associated with liposomes and aggregated on sucrose density gradients. Electron microscopy data showed that triggered F formed rosettes. Taken together these data suggest that release and membrane insertion of the hydrophobic fusion peptide require both cleavage of F and heat. Heating of cleaved F causes conversion to a postfusion form as judged by its "golf tee" morphology in the electron microscope. Heating of uncleaved F also causes conversion of F to a morphologically similar form. The reactivity of the F protein with conformation-specific mAbs and peptide binding suggest that soluble F-GCNt and membrane-bound F proteins refold through a comparable pathway.
Connolly
Sarah A
SA
Howard Hughes Medical Institute, Northwestern University, Evanston, IL 60208-3500, USA.
Leser
George P
GP
Yin
Hsien-Shen
HS
Jardetzky
Theodore S
TS
Lamb
Robert A
RA
D000818:Animals; D000911:Antibodies, Monoclonal / Q000378:metabolism; D006358:Hot Temperature; D008081:Liposomes / Q000378:metabolism*; D008854:Microscopy, Electron; D008958:Models, Molecular; D010252:Paramyxoviridae / Q000737:chemistry; D010455:Peptides / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D014760:Viral Fusion Proteins / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure*; D053586:Virus Internalization
D000911:Antibodies, Monoclonal; D008081:Liposomes; D010455:Peptides; D014760:Viral Fusion Proteins