A hyperstabilizing mutation in the base of the Ebola virus glycoprotein acts at multiple steps to abrogate viral entry

J. Maximilian Fels, Jennifer S. Spence, Robert H. Bortz, Zachary A. Bornholdt, Kartik Chandran

Research output: Contribution to journalArticlepeer-review

9 Scopus citations


Ebola virus (EBOV) causes highly lethal disease outbreaks against which no FDA-approved countermeasures are available. Although many host factors exploited by EBOV for cell entry have been identified, including host cell surface phosphatidylserine receptors, endosomal cysteine proteases, and the lysosomal cholesterol trafficking protein NPC1, key questions remain. Specifically, late entry steps culminating in viral membrane fusion remain enigmatic. Here, we investigated a set of glycoprotein (GP) mutants previously hypothesized to be entry defective and identified one mutation, R64A, that abolished infection with no apparent impact on GP expression, folding, or viral incorporation. R64A profoundly thermostabilized EBOV GP and rendered it highly resistant to proteolysis in vitro. Forward-genetics and cell entry studies strongly suggested that R64A’s effects on GP thermostability and proteolysis arrest viral entry at least at two distinct steps: The first upstream of NPC1 binding and the second at a late entry step downstream of fusion activation. Concordantly, toremifene, a small-molecule entry inhibitor previously shown to bind and destabilize GP, may selectively enhance the infectivity of viral particles bearing GP(R64A) at subinhibitory concentrations. R64A provides a valuable tool to further define the interplay between GP stability, proteolysis, and viral membrane fusion; to explore the rational design of stability-modulating antivirals; and to spur the development of next-generation Ebola virus vaccines with improved stability.

Original languageEnglish (US)
Article numbere01408-19
Issue number4
StatePublished - Jul 1 2019


  • Cell entry
  • Ebola
  • Ebolavirus
  • Filovirus
  • Fusion activation
  • Glycoprotein
  • Thermostability
  • Viral entry
  • Viral membrane fusion

ASJC Scopus subject areas

  • Microbiology
  • Virology


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