Abstract
To develop the budding yeast Pichia pastoris (Pp) as a model system for the study of protein secretion, we have characterized a secreted acid phosphatase (Pholp) from this yeast. Pholp can be induced 100-fold by starvation for phosphate. The enzyme was purified to homogeneity from a cell-wall extract by DEAE-Sepharose chromatography. We selected mutants that lacked extracellular phosphatase activity and the gene (PHO1) encoding Pholp was isolated from a recombinant plasmid library of Pp DNA by complementation of the mutant defect. PHOI encodes a protein of 468 amino acids (aa) with homology to repressible acid phosphatases from other yeast species. The sequence contains a 15-aa N-terminal signal sequence and six potential N-linked glycosylation sites. Antiserum to Pholp was used to show that Pholp transits the Pp secretory pathway in less than 5 min.
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