Intracellular maturation and secretion of acid phosphatase of Saccharomyces cerevisiae.
To elucidate intracellular maturation and secretion of acid phosphatase of Saccharomyces cerevisiae we prepared a monoclonal antibody that recognizes specifically the protein moiety of this cell surface glycoprotein. With this antibody membranes and soluble fractions of wild-type cells, grown in low-phosphate medium in the presence and absence of tunicamycin, were examined by the immunoblot technique. Similarly, secretory mutants, blocked at distinct steps in the secretory pathway at the restrictive temperature as well as a strain harboring several copies of the structural gene PHO5 for repressible acid phosphatase, were analyzed. The data suggest the following sequence of events in acid phosphatase maturation and secretion: three unglycosylated precursors with molecular masses of 60 kDa, 58 kDa and 56 kDa are synthesized into membranes of the endoplasmic reticulum, where these are core glycosylated in a membrane-bound form. They appear on sodium dodecyl sulfate gels as bands with molecular masses of 76 kDa and 80 kDa. Owing to a rate-limiting maturation step, occurring after core glycosylation, they can accumulate in a membrane-bound form. At the Golgi apparatus outer carbohydrate chains are attached to the core and the enzyme appears in a soluble form, indicating a release of acid phosphatase from the membrane between the endoplasmic reticulum and the Golgi. Pulse-chase experiments suggest that the time for acid phosphatase synthesis and its transport to the Golgi is about 5 min.
D000135:Acid Phosphatase / Q000378:metabolism*; D000911:Antibodies, Monoclonal; D001692:Biological Transport; D002462:Cell Membrane / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D006056:Golgi Apparatus / Q000201:enzymology; D007120:Immunochemistry; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D014415:Tunicamycin / Q000494:pharmacology
D000911:Antibodies, Monoclonal; D014415:Tunicamycin; D000135:Acid Phosphatase