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

Brenda L. Fredericksen, Michael Whitt

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39 Citations (Scopus)

Abstract

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, D137-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.

Original languageEnglish (US)
Pages (from-to)49-57
Number of pages9
JournalVirology
Volume217
Issue number1
DOIs
StatePublished - Mar 1 1996

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Acidic Amino Acids
Vesicular Stomatitis
Membrane Fusion
Glycoproteins
Viruses
Mutation
Mutant Proteins
Amino Acid Substitution
GTP-Binding Proteins
Rhabdoviridae
Virus Assembly
Virion
Serine
Complementary DNA

All Science Journal Classification (ASJC) codes

  • Virology

Cite this

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abstract = "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, D137-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.",
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AB - 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, D137-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.

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