title: The mod A mutant of Dictyostelium discoideum is missing the alpha 1,3-glucosidase involved in asparagine-linked oligosaccharide processing.
abstract: The recessive mutation, mod A, in the Dictyostelium discoideum strain M31 results in an alteration in the post-translational modification of lysosomal enzymes. We now report studies which indicate that mod A is deficient in glucosidase II, an enzyme which is involved in the processing of asparagine-linked oligosaccharides. [2-3H]Mannose-labeled glycopeptides were prepared from three purified mod A lysosomal enzymes and compared to the equivalent glycopeptides from parental enzymes. The mod A glycopeptides were deficient in high mannose oligosaccharides containing two phosphomannosyl residues and accumulated oligosaccharides with one phosphomannosyl residue. The phosphate was present in the form of an acid-stable phosphodiester in both instances. There was also an increase in the amount of nonphosphorylated high mannose oligosaccharides mod A and these were larger than the corresponding material from the parental enzymes. In addition, the nonphosphorylated oligosaccharides were only partially degraded by alpha-mannosidase, indicating the presence of a blocking moiety. In vitro enzyme assays demonstrated that the mod A cells cannot remove the inner 1 leads to 3-linked glucose from a glucosylated high mannose oligosaccharide. The cells are also deficient in membrane-bound neutral p-nitrophenyl-alpha-D-glucosidase activity. This activity has been attributed to glucosidase II in other systems. Removal of the outer 1 leads to 2-linked glucose from Glc3Man9Glc-NAc2 is normal, demonstrating the presence of glucosidase I activity. We conclude from these data that M31 cells are deficient in glucosidase II, the enzyme which removes the two inner glucose residues from the glucosylated oligosaccharides of newly glycosylated proteins. This defect can explain the mod A phenotype and is proposed to be the primary genetic defect in these cells.
mesh_terms: D001216:Asparagine; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004023:Dictyostelium / Q000235:genetics*; D005959:Glucosidases / Q000378:metabolism*; D008247:Lysosomes / Q000201:enzymology; D009154:Mutation; D009844:Oligosaccharides / Q000096:biosynthesis*; D010766:Phosphorylation; D013431:Sulfates / Q000378:metabolism; D000520:alpha-Glucosidases / Q000378:metabolism*
chemical_list: D009844:Oligosaccharides; D013431:Sulfates; D001216:Asparagine; D005959:Glucosidases; D000520:alpha-Glucosidases