Purification and characterization of cathepsin H fromhepatopancreas of carp Cyprinus carpio

Abstract

1. 1. Cathepsin H was purified about 5400-fold from hepatopancreas of carp Cyprinus carpio) by the method involving ammonium sulfate fractionation, and chromatography on S-sepharose, DEAE-Sephacel, Ultrogel AcA54, Concanavalin A-Sepharose 4B and GPC on Protein-Pak 125. 2. 2. The purified cathepsin H gave a single protein band on analytical-PAGE, but migrated as two bands of 27,000 and 23,000 mol. wt on SDS-PAGE. 3. 3. Cathepsin H had a pH and temperature optimum of 6.5 and 45°C using Arg-MCA as a substrate, respectively, and was activatedby sulfhydryl compounds and inhibited by cysteine protease inhibitors and metal compounds having high reactivities at cysteine residue. 4. 4. The carp hepatopancreas cathepsin H immunoreacted with the monospecific antibody against rat liver cathepsin H, and did not react with the antibodies against carp hepatopancreas cathepsin B and L by the method of immunoelectrophoretic blotting.