Mutation in Spike Protein Cleavage Site and Pathogenesis of Feline Coronavirus
Figure 4. . Furin cleavage assays of fluorogenic peptides. A) Synthetic fluorogenic peptides were generated with sequences matching consensus feline enteric coronavirus and a panel of modified sequences with substitutions (shown in red) found by feline infectious peritonitis virus sequencing. Peptides (50 μmol/L) were subjected to cleavage by recombinant human furin (1 U/100 μL), at pH 7.5, 30°C, and the release of fluorescence over time was measured by a spectrofluorometer enabling calculation of the Vmax of each reaction. Peptide cleavage scores generated by the ProP 1.0 server (www.cbs.dtu.dk/services/ProP/) are also displayed. B) For each modified peptide (substitutions shown in red), the percentage of cleavage rate compared with the canonical sequence was calculated and displayed. Cleavage assays were performed in >3 independent experiments. Error bars indicate SD for each measurement.