1. 1. A trypsin-like enzyme from the midgut gland of the Atlantic blue crab, Callinectes sapidus, was purified and studied. 2. 2. Purification was achieved by a combination of molecular sieving, ion exchange, and hydrophobic chromatography. Degree of purity was assessed by SDS-polyacrylamide electrophoresis. 3. 3. Using N-p-tosyl-l-arginine methyl ester (TAME) as a substrate, the enzyme displays optimal activity in the absence of calcium at pH 8.2. 4. 4. The tryptic activity is extremely stable between pH 7 and 8.5, but is rapidly inactivated below pH 6. Thirty-minute incubations above 50°C also inactivate the enzyme. 5. 5. The tryptic activity has a molecular weight of 33,500 da and an isoelectric point of approximately 4. 6. 6. The trypsin-like protease from Callinectes sapidus is similar to crayfish and bovine trypsins in that it is inhibited by phenylmethane sulfonylfluoride, tosyl-l-lysine chloromethyl ketone, and soybean trypsin inhibitor. Its pH optimum and its specificity for TAME and N-p-tosyl-l-lysine methyl ester (TLME) are also similar. It is concluded that the enzyme characterized in this study is trypsin.
|Number of pages||6|
|Journal||Comparative Biochemistry and Physiology - B Biochemistry and Molecular Biology|
|State||Published - 1990|
ASJC Scopus subject areas
- Molecular Biology