Formation and characterization of the trimeric form of the fusion protein of Semliki Forest virus

Enveloped animal viruses infect cells via fusion of the viral membrane with a host cell membrane. Fusion is mediated by a viral envelope glycoprotein, which for a number of enveloped animal viruses rearranges itself during fu sion to form a trimeric cu-helical coiled-coil structure, This conformation al change from the metastable, nonfusogenic form of the spike protein to th e highly stable form involved in fusion can be induced by physiological act ivators of virus fusion and also by a variety of destabilizing conditions. The E1 spike protein subunit of Semliki Forest virus (SFV) triggers membran e fusion upon exposure to mildly acidic pH and forms a homotrimer that appe ars necessary for fusion. We have here demonstrated that formation of the E 1 homotrimer was efficiently triggered under low-pH conditions but not by p erturbants such as heat or urea, despite their induction of generalized con formational changes in the E1 and E2 subunits and partial exposure of an ac id-specific E1 epitope, We used a sensitive fluorescence assay to show that neither heat nor urea treatment triggered SFV-liposome fusion at neutral p H, although either treatment inactivated subsequent low-pH-triggered fusion activity. Once formed, the low-pH-induced E1 homotrimer was very stable an d mas only dissociated under harsh conditions such as heating in sodium dod ecyl sulfate. Taken together, these data, as well as protein structure pred ictions, suggest a model in which the less stable native E1 subunit specifi cally responds to low pH to form the more stable E1 homotrimer via conforma tional changes different from those of the coiled-coil type of fusion prote ins.