D-galactosyltransferase and its endogenous substrates in chick embryo fibroblasts transformed by Rous sarcoma virus.
UDP-D-galactose: 2-acetamido-2-deoxy-beta-D-glucopyranosyl 4-beta-D-galactosyltransferase (GalTase) activity was purified, from primary chick embryo fibroblast (CEF) transformed by a temperature-sensitive, Rous sarcoma virus mutant (CEF-RSV), by chromatography on an affinity resin prepared with monoclonal antibodies to GalTase. Cellular glycopeptides from CEF, as well as CEF-RSV, maintained at permissive (37 degrees) [CEF-RSF (37 degrees)] and nonpermissive temperatures (41 degrees) [CEF-RSV (41 degrees)], were solubilized and galactosylated in vitro by incubation with purified GalTase substrates, composed of at least six discrete complex glycopeptides having bi- to tetra-antennary structures. The glycopeptides isolated from transformed cells, CEF-RSV (37 degrees), included the six types observed in nontransformed cells, but demonstrated alterations in their relative amounts, including an increase in the content of a glycopeptide containing 3 mannose and 4 glucosamine residues. Furthermore, two additional complex-type glycopeptides were isolated from CEF- but demonstrated alterations in their relative amounts, including an increase in the content of a glycopeptide containing 3 mannose and 4 glucosamine residues. Furthermore, two additional complex type glycopeptides were isolated from CEF-RSV (37 degrees). These malignant transformation-related glycopeptides were partially characterized and found to represent tri- and tetra-antennary complex glycopeptides. Endogenous galactosylation appeared to have occurred in a branched, nonspecific manner in these transformed cell-derived glycopeptides. These findings indicate that transformed cells may contain a greater preponderance of more highly branched, complex oligosaccharides which are randomly galactosylated at nonreducing termini by cellular GalTase.
D000818:Animals; D001358:Avian Sarcoma Viruses / Q000201:enzymology / Q000235:genetics*; D002241:Carbohydrates / Q000032:analysis; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002642:Chick Embryo; D005347:Fibroblasts / Q000201:enzymology; D006020:Glycopeptides / Q000302:isolation & purification; D006026:Glycoside Hydrolases; D007700:Kinetics; D007788:Lactose Synthase / Q000378:metabolism*; D009237:N-Acetyllactosamine Synthase / Q000378:metabolism*; D013379:Substrate Specificity; D013696:Temperature
D002241:Carbohydrates; D006020:Glycopeptides; D007788:Lactose Synthase; D009237:N-Acetyllactosamine Synthase; D006026:Glycoside Hydrolases