GENES FROM THE MEDICINAL LEECH (HIRUDO-MEDICINALIS) CODING FOR UNUSUAL ENZYMES THAT SPECIFICALLY CLEAVE ENDO-EPSILON(GAMMA-GLU)-LYS ISOPEPTIDE BONDS AND HELP TO DISSOLVE BLOOD-CLOTS

We previously detected in salivary gland secretions of the medicinal l eech (Hirudo medicinalis) a novel enzymatic activity, endo-epsilon(gam ma-Glu)-Lys isopeptidase, which cleaves isopeptide bonds formed by tra nsglutaminase (Factor XIIIa) between glutamine gamma-carboxamide and t he epsilon-amino group of lysine. Such isopeptide bonds, either within or between protein polypeptide chains are formed in many biological p rocesses. However, before we started our work no enzymes were known to be capable of specifically splitting isopeptide bonds in proteins. Th e isopeptidase activity we detected was specific for isopeptide bonds. The enzyme was termed destabilase. Here we report the first purificat ion of destabilase, part of its amino acid sequence, isolation and seq uencing of two related cDNAs derived from the gene family that encodes destabilase proteins, and the detection of isopeptidase activity enco ded by one of these cDNAs cloned in a baculovirus expression vector. T he deduced mature protein products of these cDNAs contain 115 and 116 amino acid residues, including 14 highly conserved Cys residues, and a re formed from precursors containing specific leader peptides. No homo logous sequences were found in public databases.