Mutations at two conserved acidic amino acids in the glycoprotein of vesicular stomatitis virus affect pH-dependent conformational changes and reduce the pH threshold for membrane fusion.
Recently, we had shown that amino acid substitutions at residues 124, 127, and 133 either abolished or drastically altered the fusion activity of the glycoprotein (G protein) of vesicular stomatitis virus (VSV), indicating that this region is important for membrane fusion and may constitute an internal fusion domain. In this report we show that amino acid substitutions at two conserved acidic residues located at the C-terminal end of the putative fusion domain also affect the fusion activity of G protein. Two substitutions, D127-L and E139-L, slightly reduced the pH threshold at which G protein mediates membrane fusion. However, both D137-L and E139-L had fusion activities equivalent to that of wild-type G protein after exposure to a pH of 5.7 or below. To determine if the fusion activity of G protein required an acidic residue in this region both D137 and E139 were replaced with serine. This double substitution also reduced the pH threshold for fusion activity, but the effect appeared to be less severe than that for either of the single substitutions. The mutated G protein cDNAs were also introduced into a VSV minigenome to examine the effect of the substitutions on virus assembly and infectivity. All three mutant G proteins assembled into particles and these virions were infectious despite the altered fusion activities of the mutant G proteins. Although particles containing the mutant proteins were infectious they appeared to be attenuated, suggesting that these two acidic residues, which are conserved in several distantly related rhabdoviruses, play an important role in maintaining virus fitness.
Fredericksen
B L
BL
Department of Microbiology and Immunology, University of Tennessee at Memphis, 38163, USA.
Whitt
M A
MA
D000818:Animals; D001665:Binding Sites; D001692:Biological Transport; D002460:Cell Line; D006224:Cricetinae; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008561:Membrane Fusion; D008562:Membrane Glycoproteins; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D011487:Protein Conformation; D014357:Trypsin / Q000378:metabolism; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism*
C028391:G protein, vesicular stomatitis virus; D008562:Membrane Glycoproteins; D014759:Viral Envelope Proteins; D014357:Trypsin