EXPRESSION, GLYCOSYLATION AND SECRETION OF YEAST ACID-PHOSPHATASE IN HAMSTER BHK-CELLS

The gene PHO5 coding for one of the repressible acid phosphatases of t he yeast Saccharomyces cerevisiae has been expressed at high efficienc y in the baby hamster kidney (BHK) cell line. The expression vector wa s constructed from PHO5 driven by the human beta-actin promoter and wa s transfected into BHK cells by the calcium phosphate method. The reco mbinant APase (r-APase) which was secreted in active form from the cel ls was estimated by SDS/polyacrylamide gel electrophoresis to have mol ecular 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 P ronase showed that the glycans are predominantly of the 2.2.4 trianten nary and tetraantennary complex-type. These data suggest that the exte nsive glycosylation of yeast APase, which contains eight polymannose s ubstiuents, is not essential for secretion and expression of enzymatic activity of the transfected gene product.