Purification and Characterization of a Trypsin‐Like Protease from Flatfish (Paralichthys olivaceus) Intestine

A trypsin‐like protease was purified from the intestine of flatfish (Paralichthys olivaceus) by gel filtration and anion‐exchange chromatography. The molecular weight was estimated to be 29.6 kDa by sodium dodecyl sulfate–polyacrylamide gel electrophoresis. Flatfish protease had maximal activity at 70C and pH 7.5 using N‐α‐benzoyl‐dl ‐arginine‐ρ‐nitroanilide as substrate. It was stable to heat treatment up to 50C and to pH ranges between 7.0 and 10.0. It was activated by calcium ion and completely inhibited by mercury ion and known serine‐protease inhibitors, such as phenylmethylsulfonyl fluoride, tosyl lysine chloromethyl ketone and benzamidine.