Mass Spectrometric Characterization of Proteins from the SARS Virus: A Preliminary Report

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

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.