Use of a monoclonal antibody to distinguish between precursor and mature forms of human lysosomal alpha-glucosidase.
The maturation of lysosomal alpha-glucosidase in cultured human skin fibroblasts was studied using a monoclonal antibody that distinguishes between the precursor and mature forms of the enzyme. Monoclonal antibodies against alpha-glucosidase isolated from placenta were produced by the hybridoma technique [Hilkens et al. (1981) Biochim. Biophys. Acta 678, 7-11]. One of these monoclonal antibodies, that synthesized by clone 43G8, reacts with the mature forms, but not with the precursor form of alpha-glucosidase isolated from urine. By means of pulse-labelling studies, it could be shown that monoclonal antibody 43G8 does not react with either the intracellular or the secreted precursor of alpha-glucosidase from cultured human skin fibroblasts. However, the antibody does react with the intermediate and mature forms of alpha-glucosidase. Endocytosis of the precurosor of alpha-glucosidase from urine by fibroblasts is followed by its conversion to a form with lower molecular mass. After endocytosis urinary precursor alpha-glucosidase is converted to a form that binds to monoclonal antibody 43G8. The t 1/2 for this conversion is 2 h. The conversion is inhibited by addition of leupeptin to the culture medium. It is concluded that a thiol proteinase is involved in the maturation of alpha-glucosidase in fibroblasts and the appearance of the antigenic determinant for 43G8.
D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D002478:Cells, Cultured; D004792:Enzyme Precursors / Q000032:analysis* / Q000276:immunology; D005347:Fibroblasts / Q000201:enzymology; D005959:Glucosidases / Q000032:analysis*; D006801:Humans; D007120:Immunochemistry; D008247:Lysosomes / Q000201:enzymology*; D012867:Skin / Q000201:enzymology; D000520:alpha-Glucosidases / Q000032:analysis* / Q000276:immunology
D000911:Antibodies, Monoclonal; D004792:Enzyme Precursors; D005959:Glucosidases; D000520:alpha-Glucosidases