Characteristics of trypsin from the pyloric ceca of walleye pollock (Theragra chalcogramma)

Trypsin was purified from the pyloric ceca of walleye pollock (Theragra chalcogramma) by gel filtration on Sephacryl S-200 and Sephadex G-50. The final enzyme preparation was nearly homogeneous in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and the molecular mass of the enzyme was estimated to be 24 kDa by SIDS-PAGE. Trypsin activity was effectively inhibited by serine protease inhibitors, such as soybean trypsin inhibitor and TLCK. Trypsin had maximal activities at around pH 8.0 and 50 degrees C for the hydrolysis of N-alpha-p-tosyl-L-arginine methyl ester hydrochloride. Trypsin was unstable above 30 degrees C and below pH 5.0, and was stabilized by calcium ions. Walleye pollock trypsin was more thermally unstable than trypsin from the Temperate Zone fish and Tropical Zone fish. The N-terminal amino acid sequence of the trypsin, IVGGYECTKHSQAHQVSLNS, was found, and the sequential identity between the walleye pollock trypsin and Frigid Zone fish trypsin was higher (85-100%) than with Temperate Zone fish trypsin (75-90%), Tropical Zone fish trypsin (75-85%), or mammalian trypsin (60-65%). (c) 2007 Elsevier Ltd. All rights reserved.