PHENOTYPIC ANALYSIS OF PROTEINASE-A MUTANTS - IMPLICATIONS FOR AUTOACTIVATION AND THE MATURATION PATHWAY OF THE VACUOLAR HYDROLASES OF SACCHAROMYCES-CEREVISIAE

We have isolated a number of mutants deficient in activity of the vacu olar hydrolase proteinase A (PrA). The mutations were sequenced and al though they all map in the PEP4 gene, which encodes the precursor to P rA, three distinguishable phenotypes have surfaced. The properties of the pep4-7 missense mutant suggested that the activation of the precur sor to proteinase A is due to an autocatalytic cleavage. PrA active si te mutations were constructed and resulted in accumulation of PrA anti gen in the inactive precursor form. Although protease B (PrB), another vacuolar hydrolase, is not required for the production of active PrA, the active form of PrA that accumulates in a strain lacking PrB is la rger than that found in a strain containing active PrB. We have purifi ed this larger form of PrA and determined that it bears 7 additional a mino acids at its NH2 terminus. It has become apparent from all the st udies performed on the maturation pathway of the vacuolar hydrolases t hat there is a great deal of redundancy built into the system.