PROTEOLYTIC CLEAVAGE OF SOYBEAN BOWMAN-BIRK INHIBITOR MONITORED BY MEANS OF HIGH-PERFORMANCE CAPILLARY ELECTROPHORESIS - IMPLICATIONS FOR THE MECHANISM OF PROTEINASE-INHIBITORS

The hydrolysis of the soybean Bowman-Birk inhibitor in the presence of catalytic amounts of bovine trypsin and the formation of the non-cova lent enzyme-inhibitor complex with an equimolar amount of enzyme are m onitored by means of high-performance capillary electrophoresis (HPCE) , The inhibitor is cleaved in the trypsin-reactive and mon slowly in t he chymotrypsin-reactive subdomain. HPCE proves itself as the only rel iable analytical tool to monitor these reactions in clear contrast to classical electrophoretic, chromatographic and enzymatic methods. The most efficient separation of the intact and the two active site cleave d forms of the inhibitor was achieved in berate buffer at pH 10.0. The pH dependence of the rate constant and the final extent of hydrolysis reveal the stability of the enzyme inhibitor complex as a central asp ect of the mechanism of proteinase inhibitors.