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Originally published In Press as doi:10.1074/mcp.M300048-MCP200 on May 29, 2003.
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Molecular & Cellular Proteomics 2:346-356, 2003.
© 2003 by The American Society for Biochemistry and Molecular Biology, Inc.

Research

Mass Spectrometric Characterization of Proteins from the SARS Virus

A Preliminary Report*,S

Oleg Krokhin{ddagger},§,¶, Yan Li||,¶, Anton Andonov||, Heinz Feldmann||,**, Ramon Flick||,**, Steven Jones||, Ute Stroeher||,**, Nathalie Bastien||, Kumar V. N. Dasuri§, Keding Cheng§, J. Neil Simonsen**, Hélène Perreault{ddagger}{ddagger}, John Wilkins§, Werner Ens{ddagger},§, Frank Plummer||,§§ and Kenneth G. Standing{ddagger},§,§§

{ddagger} Physics and Astronomy, University of Manitoba, Winnipeg MB R3T 2N2, Canada
§ Manitoba Centre for Proteomics, 805 JBRC, 715 McDermot Avenue, Winnipeg MB R3E 3P4, Canada
|| National Microbiology Laboratory, 1015 Arlington Street, Winnipeg MB R3E 3R2, Canada
** Department of Medical Microbiology, University of Manitoba, 730 William Avenue, Winnipeg MB R3E OW3, Canada
{ddagger}{ddagger} Department of Chemistry, University of Manitoba, Winnipeg MB R3T 2N2, Canada

A new coronavirus has been implicated as the causative agent of severe acute respiratory syndrome (SARS). We have used convalescent sera from several SARS patients to detect proteins in the culture supernatants from cells exposed to lavage another SARS patient. The most prominent protein in the supernatant was identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) as a ~46-kDa species. This was found to be a novel nucleocapsid protein that matched almost exactly one predicted by an open reading frame in the recently published nucleotide sequence of the same virus isolate (>96% coverage). A second viral protein corresponding to the predicted ~139-kDa spike glycoprotein has also been examined by MALDI-TOF MS (42% coverage). After peptide N-glycosidase F digestion, 12 glycosylation sites in this protein were confirmed. The sugars attached to four of the sites were also identified. These results suggest that the nucleocapsid protein is a major immunogen that may be useful for early diagnostics, and that the spike glycoprotein may present a particularly attractive target for prophylactic intervention in combating SARS.


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