Plasmin cleaves HK to release BK in buffer. (A) Western blot depicting cleavage pattern of purified hHK enzymatically processed in vitro. The enzymatic concentration of PKa or plasmin, or vehicle (veh) control, is indicated below each lane, and incubation proceeded for 1 hour at 37°C. (B) Concentration of BK released in an enzymatic buffer system, measured by using ELISA. HK 80 µg/mL was incubated for 1 hour with either PKa or plasmin at indicated concentrations. Error bars represent standard deviation from the mean. (C) Mass spectra of BK standard (250 µg/mL). (D) Mass spectra of purified HK (800 µg/mL). (E) Mass spectra of HK (800 µg/mL) incubated with pKA (150 µg/mL) for 1 hour. (F) Mass spectra of HK (800 µg/mL) incubated with plasmin (150 µg/mL) for 1 hour. Molecular mass of 1060 interpreted as BK peptide.