CYSTATIN-LIKE DOMAIN OF MOUSE COUNTERTRYPIN, A MEMBER OF MAMMALIAN FETUIN FAMILY, IS RESPONSIBLE FOR THE INHIBITION OF TRYPSIN - EVIDENCE FROM SITE-DIRECTED MUTAGENESIS

Members of mammaliar, fetuin family, such as human alpha(2)HS glycopro tein and bovine fetuin, consist of three domains, two tandemly arrange d cystatin-like domains and an unrelated domain, but they have no inhi bitory activity against cysteine proteinases. We found that countertry pin, a novel trypsin inhibitor, is mouse counterpart of human alpha(2) HS glycoprotein, and that human alpha(2)HS glycoprotein and bovine fet uin which were prepared without use of ethanol are capable of inhibiti ng trypsin (Yamamoto, K. and Sinohara, H. (1993) J. Biol. Chem, 268, 1 7750-17753). In the present study, cDNA encoding countertrypin was iso lated and sequenced, and evidence is presented, based on the site-dire cted mutagenesis, that lysine-231 in the second cystatin domain is the P1 site for trypsin inhibition.