title: Beta-galactosidase-deficient human fibroblasts: uptake and processing of the exogenous precursor enzyme expressed by stable transformant COS cells.
abstract: COS-1 cells were transfected by electroporation with a cDNA for human acid beta-galactosidase cloned in our laboratory and stable transformants expressing the enzyme activity were selected. The precursor form of the enzyme was secreted in large quantities into the culture medium. The fibroblasts from patients with GM1-gangliosidosis or Morquio B disease showed a remarkable increase of enzyme activity, up to the normal level, after culture in this medium for 2 days; the amount of uptake was essentially the same as that for the precursor form in human fibroblasts. After endocytosis, the precursor molecules were processed normally to the mature form and remained as stable as those produced by human fibroblasts. On the other hand, cells from galactosialidosis patients did not show any increase of enzyme activity in a similar experiment. It was concluded that the transformants are useful as the source of precursor proteins for the study of intracellular turnover of enzyme molecules in mutant cells.
authors.affiliation: Department of Clinical Genetics, Tokyo Metropolitan Institute of Medical Science, Japan.
authors.forename: A
authors.initials: A
authors.lastname: Oshima
authors.forename: K
authors.initials: K
authors.lastname: Itoh
authors.forename: Y
authors.initials: Y
authors.lastname: Nagao
authors.forename: H
authors.initials: H
authors.lastname: Sakuraba
authors.forename: Y
authors.initials: Y
authors.lastname: Suzuki
mesh_terms: D000818:Animals; D002268:Carboxypeptidases / Q000235:genetics; D043402:Cathepsin A; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D004705:Endocytosis; D004792:Enzyme Precursors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology*; D005733:Gangliosidoses / Q000201:enzymology; D005822:Genetic Vectors / Q000235:genetics; D006023:Glycoproteins / Q000235:genetics; D006801:Humans; D009085:Mucopolysaccharidosis IV / Q000201:enzymology; D014162:Transfection / Q000235:genetics; D001616:beta-Galactosidase / Q000172:deficiency* / Q000235:genetics / Q000378:metabolism
chemical_list: D004792:Enzyme Precursors; D006023:Glycoproteins; D001616:beta-Galactosidase; D002268:Carboxypeptidases; C468629:CTSA protein, human; D043402:Cathepsin A