Compartmentalized assembly of oligosaccharides on exported glycoproteins in yeast.
Temperature-sensitive secretory mutants (sec) of S. cerevisiae have been used to evaluate the stages and localization of glycoprotein oligosaccharide synthesis. At the nonpermissive growth temperature (37 degrees C), the sec mutants accumulate secretory organelles and glycoproteins. Histochemical staining and thin-section electron microscopy reveal that the secreted glycoprotein, acid phosphatase, is contained within one of three distinct organelles that accumulates in different mutants: ER; Golgi-like structures called Berkeley bodies; and 80--100 nm vesicles. When produced at 37 degrees C, invertase and acid phosphatase have less carbohydrate in the mutants that accumulate ER than in other mutants, or than in the wild-type strain. External invertase migrates on SDS-polyacrylamide gels as a heterogeneous species with an apparent molecular weight of 100 to 140 kd. Radiolabeled invertase, immunoprecipitated from extracts of ER-accumulating mutant cells, migrates as a set of three discrete protein species with apparent molecular weights of 79, 81, and 83 kd; the other mutants produce a form more like the secreted enzyme. In each case, removal of N-glycosidically linked oligosaccharides by treatment with endoglycosidase H produces a discrete species that migrates as a protein of 61 kd. Immunochemical analysis of bulk glycoprotein accumulated in the mutants suggests that a major portion of the N-linked oligosaccharide, the outer chain, is added after material passes from the ER.
D000135:Acid Phosphatase / Q000378:metabolism; D002451:Cell Compartmentation; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000378:metabolism; D008351:Mannans / Q000096:biosynthesis; D009154:Mutation; D009844:Oligosaccharides / Q000096:biosynthesis*; D009940:Organoids / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism* / Q000648:ultrastructure; D013393:Sucrase / Q000378:metabolism; D043324:beta-Fructofuranosidase
D006023:Glycoproteins; D008351:Mannans; D009844:Oligosaccharides; D000135:Acid Phosphatase; D006026:Glycoside Hydrolases; D043324:beta-Fructofuranosidase; D013393:Sucrase