Selective release of the disaccharide 2-acetamido-2-deoxy-3-O-(beta-D-galactopyranosyl)-D-galactose from epiglycanin by endo-N-acetyl-alpha-D-galactosaminidase.
Epiglycanin, the major glycoprotein of TA3-Ha mammary carcinoma ascites cells, was radiolabeled with tritium in the terminal D-galactose and 2-acetamido-2-deoxy-D-galactose residues. Alkaline-borohydride treatment, reported to release five O-glycosyl-linked chain types from epiglycanin, resulted in the cleavage of 98-99% of the radioactivity from the protein. Of this, 63% of the radioactivity from epiglycanin and 70% from asialoepiglycanin co-migrated with an authentic sample of 2-acetamido-2-deoxy-3-O-(beta-D-galactopyranosyl)-D-galactitol on a column of Bio-Gel P-6. Incubation of [3H]galactose-epiglycanin with endo-N-acetyl -alpha-D-galactosaminidase (Diplococcus pneumoniae), and fractionation of the mixture on a column of Bio-Gel P-4, gave only one oligosaccharide peak containing 62 and 70%, respectively, of the radioactivity of epiglycanin and asialoepiglycanin. This oligosaccharide comigrated with authentic 2-acetamido-2-deoxy-3-O-(beta-D-galactopyranosyl)-D-galactose (1) on columns of Bio-Gel P-2 and P-4 and on paper chromatograms. Results of experiments in which unlabeled epiglycanin was treated with enzyme and the products analyzed, by three different methods, suggested that 78-85% of 1 had been cleaved. Another enzyme, N-acetyl-alpha-D-galactosaminyl-oligosaccharidase from Clostridium perfringens, exhibited similar specificity and cleaved 65% of the radioactivity from ([3H]galactose)asialoepiglycanin, which was eluted from a Bio-Gel P-2 column as the disaccharide 1.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D015295:Antigens, Tumor-Associated, Carbohydrate; D004187:Disaccharides / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism*; D006595:Hexosamines / Q000032:analysis; D006596:Hexosaminidases / Q000378:metabolism*; D008325:Mammary Neoplasms, Experimental / Q000378:metabolism*; D008562:Membrane Glycoproteins; D000519:alpha-Galactosidase / Q000378:metabolism; D048809:alpha-N-Acetylgalactosaminidase
D000596:Amino Acids; D015295:Antigens, Tumor-Associated, Carbohydrate; D004187:Disaccharides; D006023:Glycoproteins; D006595:Hexosamines; D008562:Membrane Glycoproteins; C016424:epiglycanin protein, mouse; C016467:Thomsen-Friedenreich antigen; D006596:Hexosaminidases; C039044:N-acetyl-alpha-D-galactosaminyloligosaccharidase; D000519:alpha-Galactosidase; D048809:alpha-N-Acetylgalactosaminidase