Mass spectrometry analysis of the Ebola virus soluble glycoprotein sGP identified a rare post-translation modification, C-mannosylation, which was found on tryptophan (W) 288. This modification has not been described for any other viral protein; however, many viral transmembrane glycoproteins contain one or more of the recognition motifs (W-x-x-W). Elimination of the C-mannose on sGP did not significantly alter protein biosynthesis, processing or structure. Furthermore, the protective effect of sGP on endothelial barrier function, currently the only known activity of sGP, was unaltered. It is possible that C-mannosylation may be a common post-translational modification of viral transmembrane glycoproteins where it could play a role in particle maturation and/or entry by stabilizing the structure of these proteins. In this regard, C-mannosylation of sGP may be an anomaly resulting from the unique manner in which this protein is generated as the product of unedited transcripts from the glycoprotein gene of Ebola.