Biochemical evidence of specific trypsin-chymotrypsin inhibitors in the rhynchobdellid leech, Theromyzon tessulatum

The presence of two specific trypsin-chymotrypsin inhibitors from head part s of the rhynchobdellid leech Theromyzon tessulatum is reported. Two protei ns, anti-trypsin-chymotrypsin A (ATCA; 14636.6 +/- 131 Dal and anti-trypsin -chymotrypsin B (ATCB; 14368 +/- 95 Dal were purified by size exclusion and anion-exchange chromatography followed by reversed-phase HPLC. Based on am ino-acid composition, N-terminal sequence determination (MELCELGQSCSRDNPQPS NM), matrix assisted laser desorption-time of flight measurement (MALDI-TOF ), trypsin mapping comparison, inhibition constant determination (K-i), and influence on amidolytic activity of different serine proteases, it is demo nstrated that ATCA and ATCB are novel and highly potent serine-protease inh ibitors of trypsin and chymotrypsin (ATCA: 350 fM towards trypsin and chymo trypsin; ATCB: 400 and 75 fM towards trypsin and chymotrypsin, respectively ). It is further surmised that ATCA and ATCB are linked, in that ATCB would lead to the formation of ATCA after loss of few amino acid residues.