ISOLATION AND CHARACTERIZATION OF A TRYPSIN FROM THE SLIPPER LOBSTER,THENUS-ORIENTALIS (LUND)

Trypsin has been isolated and purified from the digestive glands of th e slipper lobster, Thenus orientalis. It is a glycoprotein with a mole cular mass of approximately 35 kDa as judged by both SDS-PAGE and gel filtration. The N-terminal amino acid sequence has strong homology to crustacean trypsins. This is confirmed by the cross-reaction of crusta cean trypsins with antibodies to the T. orientalis enzyme, Despite a 4 0% identity with the bovine trypsin N-terminal sequence, there was no cross-reaction with the mammalian serine proteases. The optimum k(cat) and k(cat)/K-m values for N-alpha-benzoylarsnine-p-nitroanalide were 0.91 s(-1) and 9.7 X 10(3) M(-1) s(-1), respectively, with this specif icity constant being lower than those reported for other crustacean tr ypsins, Inhibition studies indicated the presence of serine and histid ine at the active site and pK(a)lpha of the catalytic histidine residu e was found to be 5.7 in the free enzyme and 4.7 in the Michaelis comp lex. (C) 1995 Academic Press, Inc.