A NOVEL MEMBRANE-ASSOCIATED METALLOPROTEASE, STE24P, IS REQUIRED FOR THE FIRST STEP OF NH2-TERMINAL PROCESSING OF THE YEAST A-FACTOR PRECURSOR

Many secreted bioactive signaling molecules, including the yeast matin g pheromones a-factor and alpha-factor, are initially synthesized as p recursors requiring multiple intracellular processing enzymes to gener ate their mature forms. To identify new gene products involved in the biogenesis of a-factor in Saccharomyces cerevisiae, we carried out a s creen for MA Ta-specific, mating-defective mutants. We have identified a new mutant, ste24, in addition to previously known sterile mutants. During its biogenesis in a wild-type strain, the a-factor precursor u ndergoes a series of COOH-terminal CAAX modifications, two sequential NH2-terminal cleavage events, and export from the cell. Identification of the a-factor biosynthetic intermediate that accumulates in the ste 24 mutant revealed that STE24 is required for the first NH2-terminal p roteolytic processing event within the a-factor precursor, which takes place after COOH-terminal CAAX modification is complete. The STE24 ge ne product contains multiple predicted membrane spans, a zinc metallop rotease motif (HEXXH), and a COOH-terminal ER retrieval signal (KKXX). The HEXXH protease motif is critical for STE24 activity, since STE24 fails to function when conserved residues within this motif are mutate d. The identification of Ste24p homologues in a diverse group of organ isms, including Escherichia coli, Schizosaccharomyces pombe, Haemophil us influenzae, and Homo sapiens, indicates that Ste24p has been highly conserved throughout evolution. Ste24p and the proteins related to it define a new subfamily of proteins that are likely to function as int racellular, membrane-associated zinc metalloproteases.