Purification and characterization of serine protease inhibitor from white croaker Argyrosomus argentatus ordinary muscle.

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Abstract
A serine protease inhibitor was isolated from white croaker Argyrosomus argentatus ordinary muscle by ammonium sulfate fractionation, gel filtration on Sephadex G-100, column chromato-graphies on CM-Sephadex C-50 and DEAE-Sephadex A-50, and preparative polyacrylamide gel electrophoresis. The inhibitory activity against trypsin increased about 1, 500-fold, and the purified inhibitor was homogeneous as judged by polyacrylamide gel electrophoresis. The molecular weight of the inhibitor was estimated to be about 100, 000 by gel filtration on Sephadex G-100, but was estimated to be about 55, 000 by SDS polyacrylamide gel electrophoresis after reduction with 2-mercaptoethanol. The inhibitor was stable over a range of pH 6.0-9.0, but unstable below pH 5.0, and completely inactivated by heating at 60°C for 30 min at pH 6.0. The isoelectric point was found to be about 4.7, and N-terminal glycine was found by dansylation. Trypsin, α-chymotrypsin and elastase were strongly inhibited with the inhibitor, but pepsin, thermolysin and papain were not inhibited. Each E-I complex of the inhibitor with the proteases, trypsin, α-chymotrypsin and elastase, were stable to the denaturing reagents such as SDS and 2-mercaptoethanol, however each complex was digested in the presence of the excess protease.

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