title: Expression, glycosylation and secretion of yeast acid phosphatase in hamster BHK cells.
abstract: The gene PHO5 coding for one of the repressible acid phosphatases of the yeast Saccharomyces cerevisiae has been expressed at high efficiency in the baby hamster kidney (BHK) cell line. The expression vector was constructed from PHO5 driven by the human beta-actin promoter and was transfected into BHK cells by the calcium phosphate method. The recombinant APase (r-APase) which was secreted in active form from the cells was estimated by SDS/polyacrylamide gel electrophoresis to have molecular mass M(r) = 62,000, indicating substitution of the polypeptide moiety by 2-3 asparagine-linked glycans. Analysis by sequential lectin affinity chromatography of glycopeptides obtained from r-APase with Pronase showed that the glycans are predominantly of the 2.2.4 triantennary and tetraantennary complex-type. These data suggest that the extensive glycosylation of yeast APase, which contains eight polymannose substituents, is not essential for secretion and expression of enzymatic activity of the transfected gene product.
authors.affiliation: Laboratory of Biochemistry, Faculty of Food Technology and Biotechnology, University of Zagreb, Yugoslavia.
authors.forename: R
authors.initials: R
authors.lastname: Reljic
authors.forename: S
authors.initials: S
authors.lastname: Barbaric
authors.forename: B
authors.initials: B
authors.lastname: Ries
authors.forename: R
authors.initials: R
authors.lastname: Buxton
authors.forename: R C
authors.initials: RC
authors.lastname: Hughes
mesh_terms: D000135:Acid Phosphatase / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology*; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors / Q000235:genetics; D006020:Glycopeptides / Q000032:analysis; D006031:Glycosylation; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D014170:Transformation, Genetic / Q000235:genetics
chemical_list: D006020:Glycopeptides; D000135:Acid Phosphatase