MATURATION OF HUMAN PROCATHEPSIN-B - PROENZYME ACTIVATION AND PROTEOLYTIC PROCESSING OF THE PRECURSOR TO THE MATURE PROTEINASE, IN-VITRO, ARE PRIMARILY UNIMOLECULAR PROCESSES

Recombinant latent human procathepsin B produced in yeast was purified to near homogeneity. The purified recombinant proenzyme is activated in vitro under acidic conditions resulting in rapid conversion into th e mature form of the proteinase. Activation as well as proteolytic mat uration of the recombinant cathepsin B precursor were shown to be prim arily concentration-independent processes indicating a unimolecular (i .e. intramolecular) mechanism. Only one cleavage site was identified, yielding a mature polypeptide with the same amino-terminal sequence as that found in recombinant active human cathepsin B obtained from yeas t culture media. The same peptide bond is cleaved during processing of a nonactivatable mutant of procathepsin B by the purified mature enzy me (i.e. intermolecular processing). Thus, the complete proregion is l iberated during procathepsin B processing. This peptide may then act a s a reversible inhibitor and stabilizer of the mature proteinase, and it appears likely that cathepsin B-propeptide complexes occur transien tly during proteolytic maturation.