Glycosylation and processing of carbohydrate side chains of ecto-5'-nucleotidase in cultured human chorionic cells.
Glycosylation and carbohydrate processing of ecto-5'-nucleotidase were studied in cultured human chorionic cells using metabolic labelling and immunoprecipitation with monoclonal antibodies. Tunicamycin blocks glycosylation altogether leading to a reduction in molecular mass of 9,500 Da. The same result is obtained by digesting the mature 72,000-Da protein with endoglycosidase F. Using various inhibitors of the carbohydrate-trimming reactions like deoxynojirimycin, deoxymannojirimycin and swainsonine smaller molecular mass reductions are observed and the oligosaccharide side chains are kept in a configuration sensitive to endoglycosidase H digestion. Digestion of mature 5'-nucleotidase with endoglycosidase H leads to a much smaller (2,000 Da) reduction in molecular mass. It is calculated that, in addition to the phosphatidylinositol-glycan anchor structure, ecto-5'-nucleotidase of human chorionic cells should carry 4 oligosaccharide side chains per subunit, 3 of which should be of the complex and one of the high mannose type. Interference with carbohydrate processing by various inhibitors does not seem to influence the distribution of ecto-5'-nucleotidase between the cell surface and intracellular membranes nor does it block the transfer of the enzyme to the phosphatidylinositol glycan anchor.
Burgemeister
R
R
Institut für Anthropologie und Humangenetik, Universität München.
Danescu
I
I
Gutensohn
W
W
D017485:1-Deoxynojirimycin; D015720:5'-Nucleotidase / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D000470:Alkaloids / Q000494:pharmacology; D050260:Carbohydrate Metabolism; D002478:Cells, Cultured; D002824:Chorionic Villi / Q000201:enzymology; D005944:Glucosamine / Q000031:analogs & derivatives / Q000494:pharmacology; D006031:Glycosylation; D006801:Humans; D008361:Mannosidases / Q000037:antagonists & inhibitors; D017026:Swainsonine; D014415:Tunicamycin / Q000494:pharmacology
D000470:Alkaloids; D014415:Tunicamycin; D017485:1-Deoxynojirimycin; D015720:5'-Nucleotidase; D008361:Mannosidases; D005944:Glucosamine; D017026:Swainsonine