An extractable collagenase from crustacean hepatopancreas

Abstract

An enzyme capable of degrading native collagen under physiologic conditions has been extracted from the hepatopancreas of a crustacean, Uca pugilator. The collagenase acts on native collagen fibrils and on collagen in solution. At 25° the enzyme is capable of producing a marked reduction in the specific viscosity of collagen with no loss of optical rotation, indicating the ability of this enzyme to cleave the native collagen helix without producing denaturation. Collagenolytic activity, as investigated by disc electrophoresis, results in the production of numerous new components below the original α band. Three fragments, 75% (TC A 75), 70% (TC A 70) and 67% (TC A 67) the molecular length from the “A” or N-terminal end of the molecule, have been identified in electron micrographs of segment-long-spacing crystallites prepared from enzyme-collagen reaction mixtures. Crude collagenase preparations from this digestive organ also contain significant chymotrypsin- and trypsin-like activities. The cleavage by hepatopancreas extracts of peptide bonds in the nonhelical, N-terminal region of collagen, at or near the intramolecular cross-link, may be due to the action of chymotrypsin on collagen. Chymotrypsin, however, is unable to degrade α chains further at 25°. Trypsin has little effect on the viscosity of collagen in solution at concentrations equivalent to those present in hepatopancreas extracts. At these concentrations trypsin is unable to reduce the content of cross-linked components in collagen or catalyze cleavages in the native helix. In addition, the ability of hepatopancreas extracts, but not chymotrypsin or trypsin, to degrade collagen in fibrillar form further indicates the presence of a true collagenase in crustacean hepatopancreas.