PROTEOLYSIS OF TYPE-III COLLAGEN BY COLLAGENASE AND CATHEPSIN-B UNDERHIGH HYDROSTATIC-PRESSURE

The effects of high hydrostatic pressures on the kinetics of hydrolysi s of type III collagen from calfskin by collagenase and cathepsin B we re studied. Collagen hydrolysates sampled at different time intervals (0-90 min) and at different pressures (0.1-300 MPa) were analysed by r everse-phase high pressure liquid chromatography. The rate of collagen hydrolysis decreased up to 300 MPa for both enzymes. The rate of coll agen hydrolysis with cathepsin B was drastically reduced between 0.1 a nd 100 MPa. Significant differences in the populations of the peptides in cathepsin B hydrolysates were observed in chromatograms correspond ing to different pressures. This indicated that some amino acid side-c hains were less exposed to cathepsin B recognition on the surface of c ollagen molecules at high pressures. In contrast, the chromatograms of collagenase hydrolysates showed similar patterns, varying only by the peak heights in chromatograms corresponding to the 0.1-300 MPa pressu re range. Pressure-induced decreases of the enzyme-apparent activities suggested that the activation volumes for the reaction of both enzyme s were positive.