Novel preparation of functional Sindbis virosomes.

Lipid and protein factors important for the preparation and stability of reconstituted membranes prepared by the insertion of detergent-solubilized Sindbis virus glycoproteins into preformed lipid vesicles have been defined. It was found that both the state of aggregation of the membrane protein and the phase of the lipid are critical for the insertion of proteins into preformed lipid vesicles. The membranes prepared with the insertion technique were characterized in terms of residual detergent, protein orientation, and whether or not they were sealed. Binding and fusion experiments have been carried out with the insertion membranes and virus. It was found that BHK-21 cells at 4 degrees C bound one-fifth to one-tenth the number of insertion membranes as intact virus, and binding was saturable in both cases. Variation of the lipid/protein ratio did not result in significant differences in binding. The insertion membranes were found to fuse to a model lipid bilayer at least as well as the virus. These results are discussed in terms of structural factors important for the biological functionalities of the viral spike glycoproteins.