Activation of the SARS coronavirus spike protein via sequential proteolytic cleavage at two distinct sites

Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5871-6. doi: 10.1073/pnas.0809524106. Epub 2009 Mar 24.


The coronavirus spike protein (S) plays a key role in the early steps of viral infection, with the S1 domain responsible for receptor binding and the S2 domain mediating membrane fusion. In some cases, the S protein is proteolytically cleaved at the S1-S2 boundary. In the case of the severe acute respiratory syndrome coronavirus (SARS-CoV), it has been shown that virus entry requires the endosomal protease cathepsin L; however, it was also found that infection of SARS-CoV could be strongly induced by trypsin treatment. Overall, in terms of how cleavage might activate membrane fusion, proteolytic processing of the SARS-CoV S protein remains unclear. Here, we identify a proteolytic cleavage site within the SARS-CoV S2 domain (S2', R797). Mutation of R797 specifically inhibited trypsin-dependent fusion in both cell-cell fusion and pseudovirion entry assays. We also introduced a furin cleavage site at both the S2' cleavage site within S2 793-KPTKR-797 (S2'), as well as at the junction of S1 and S2. Introduction of a furin cleavage site at the S2' position allowed trypsin-independent cell-cell fusion, which was strongly increased by the presence of a second furin cleavage site at the S1-S2 position. Taken together, these data suggest a novel priming mechanism for a viral fusion protein, with a critical proteolytic cleavage event on the SARS-CoV S protein at position 797 (S2'), acting in concert with the S1-S2 cleavage site to mediate membrane fusion and virus infectivity.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Binding Sites
  • Cell Fusion
  • Furin
  • Hydrolysis
  • Membrane Glycoproteins / metabolism*
  • Membrane Glycoproteins / physiology*
  • Mutagenesis, Site-Directed
  • SARS Virus / pathogenicity*
  • Spike Glycoprotein, Coronavirus
  • Trypsin / metabolism*
  • Viral Envelope Proteins / metabolism*
  • Viral Envelope Proteins / physiology*
  • Virus Internalization


  • Membrane Glycoproteins
  • Spike Glycoprotein, Coronavirus
  • Viral Envelope Proteins
  • spike glycoprotein, SARS-CoV
  • spike protein, mouse hepatitis virus
  • Trypsin
  • Furin