The effect of oligosaccharide chains of different sizes on the maturation and physical properties of the G protein of vesicular stomatitis virus

The envelope glycoprotein, G, of vesicular stomatitis virus (VSV) is initially glycosylated by the en bloc transfer of Glc₃Man₉GlcNAc₂ oligosaccharides to 2 specific asparagine residues in the nascent polypeptide chain. We carried out in vivo and in vitro studies to determine whether the size of the oligosaccharide chains on two related but different G proteins can affect their ability to fold correctly. For the in vivo studies we used a mutant lymphoma cell line, Thy-1^-e, which transfers the truncated oligosaccharide, Glc₃Man₅GlcNAc₂, to nascent polypeptides. The growth of VSV in these cells was temperature-sensitive compared to that in parental Thy-1⁺ cells, and VSV (San Juan) was more affected than VSV (Orsay). These results are congruous with our previous observation that in the absence of glycosylation virus assembly is temperature-sensitive and VSV (San Juan) is inhibited more than VSV (Orsay). To examine the effect of oligosaccharide size on the properties of the G protein in vitro we treated G proteins containing either Man₈GlcNAc₂ or Man₅GlcNAc₂ oligosaccharide chains with guanidine hydrochloride and measured their ability to refold using an in vitro aggregation assay. The San Juan G protein with Man₅GlcNAc₂ oligosaccharides aggregated at 40°C but not at 30°C. The Orsay G protein with Man₅GlcNAc₂ oligosaccharides and both proteins containing Man₈GlcNAc₂ oligosaccharides did not aggregate at either temperature. We conclude that the size of the oligosaccharides present on the folding G protein can be crucial in attaining a proper conformation, and the extent of their effect depends on the primary structure of the polypeptide.