Folding, unfolding, and refolding of the vesicular stomatitis virus glycoprotein.

Folding and refolding of the vesicular stomatitis virus (VSV) glycoprotein (G protein), New Jersey serotype, were studied both in infected cells and after urea denaturation and reduction of isolated protein in vitro. To assess the contribution of disulfide bonds to the conformation of this type I membrane glycoprotein, reduced and alkylated forms were compared with unreduced G proteins by their mobility on SDS−polyacrylamide gels and by their reactivity with conformation-dependent monoclonal antibodies (MAbs). Pulse−chase experiments showed that G protein folding in the endoplasmic reticulum (ER) of infected cells occurred rapidly (estimated half-time of 1−2 min) and involved transient association with the ER chaperone calnexin. Inhibition of glycosylation by tunicamycin slowed the folding process and emergence from the ER but did not prevent the appearance of a conformationally mature transport-competent G protein. For in vitro refolding studies, native G protein isolated from virus particles was denatur...