Biochemical consequences of a mutation that controls the cholesterol dependence of Semliki Forest virus fusion

Prodyot K. Chatterjee, Malini Vashishtha, Margaret Kielian

Research output: Contribution to journalArticle

45 Scopus citations

Abstract

The enveloped alphavirus Semliki Forest virus (SFV) infects cells via a low-pH-triggered membrane fusion reaction that requires cholesterol and sphingolipid in the target membrane. Cholesterol-depleted insect cells are highly resistant to alphavirus infection and were used to select srf-3, an SFV mutant that is ~100-fold less cholesterol dependent for infection due to a single amino acid change in the E1 spike subunit, proline 226 to serine. Sensitive lipid-mixing assays here demonstrated that the in vitro fusion of srf-3 and wild-type (wt) virus with cholesterol-containing liposomes had comparable kinetics, activation energies, and sphingolipid dependence. In contrast, srf-3 fusion with sterol-free liposomes was significantly more efficient than that of wt virus. Thus, the srf-3 mutation does not affect its general fusion properties with purified lipid bilayers but causes a marked and specific reduction in cholesterol dependence. Upon exposure to low pH, the E1 spike subunit undergoes distinct conformational changes, resulting in the exposure of an acid conformation-specific epitope and formation of an E1 homotrimer. These conformational changes were strongly cholesterol and sphingolipid dependent for wt SFV and strikingly less cholesterol dependent for srf-3. Our results thus demonstrate the functional importance of fusogenic E1 conformational changes in the control of SFV cholesterol dependence.

Original languageEnglish (US)
Pages (from-to)1623-1631
Number of pages9
JournalJournal of virology
Volume74
Issue number4
DOIs
StatePublished - 2000

ASJC Scopus subject areas

  • Microbiology
  • Immunology
  • Insect Science
  • Virology

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