An inducible acid phosphatase from the yeast Pichia pastoris: characterization of the gene and its product.
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 (Pho1p) from this yeast. Pho1p 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 Pho1p was isolated from a recombinant plasmid library of Pp DNA by complementation of the mutant defect. PHO1 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 Pho1p was used to show that Pho1p transits the Pp secretory pathway in less than 5 min.
Payne
W E
WE
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139, USA.
Gannon
P M
PM
Kaiser
C A
CA
D000135:Acid Phosphatase / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D001483:Base Sequence; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D004790:Enzyme Induction; D015723:Gene Library; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D005838:Genotype; D007700:Kinetics; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010843:Pichia / Q000201:enzymology* / Q000235:genetics*; D010957:Plasmids; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid
D004271:DNA, Fungal; D000135:Acid Phosphatase