Fig. 2

Purification and identification of Kn-Ba. a The third chromatographic peak (F3) obtained from the first purification step was subjected to another molecular exclusion chromatography on a Superdex 75 10/300 GL column, equilibrated and eluted with ammonium acetate 50 mM in a climate-controlled room (22 ± 2 °C). Samples were collected at a flow rate of 0.4 mL/min, and their protein content was monitored by recording the absorbance at 280 nm. b Zoom and highlight of F3–1, substrate-cleaving activity containing peak. c Silver-stained SDS-PAGE (10%) of the 33 kDa protein band corresponding to F3–1, which was denominated Kn-Ba. d The proteolytic activity of Kn-Ba, pretreated or not with EDTA, PMSF and PHE upon Abz-FRSSR-EDDnp FRET substrate. This assay was performed in quadruplicate. Results were expressed as inhibition percentage of proteolytic activity ± SEM and analyzed statistically using One-Way ANOVA test followed by Tukey HSD post-hoc tests (*p < 0.05)