ISOLATION AND COMPARATIVE PROPERTIES OF SHRIMP TRYPSIN

An enzyme with proteolytic activity was isolated from the digestive gland (hepatopancreas) of the white shrimp (Penaeus setiferus). A trypsin-like specificity was suggested on the basis of assays conducted on synthetic substrates. The enzyme was isolated by homogenizing the digestive gland with cold acetone and subjecting the resulting powder to several purification steps consisting of salt fractionation, Sephadex G-75 chromatography, and ion-exchange chromatography on DEAE-Sephadex A-50. The preparation was judged to be a single, homogeneous protein by disc electrophoresis and by ultracentrifugation. Characterization of the purified enzyme revealed a number of properties similar to mammalian trypsin. These properties included a molecular weight of 24,000 and inhibition by diisopropylphosphorofluoridate, 1-chloro-3-tosylamido-7-amino-2-heptanone, and soybean trypsin inhibitor, but not by L-tosylamido-2-phenylethylchloromethyl ketone. In contrast, shrimp trypsin has an acidic isoelectric point, no requirement for Ca2+ for stability of the enzyme, resistance to autodigestion, and irreversible inactivation below pH 5.0. In addition, the amino acid composition of the molecule suggests some differences in total structure and there seems to be no indication that the enzyme has a zymogen form. Significantly, no chymotrypsin esterase activity is present in hepatopancreas extracts, indicative of a different protein digestion pattern in this species. © 1969, American Chemical Society. All rights reserved.