Mucin synthesis. Conversion of R1-beta 1-3Gal-R2 to R1-beta 1-3(GlcNAc beta 1-6)Gal-R2 and of R1-beta 1-3GalNAc-R2 to R1-beta 1-3(GlcNAc beta 1-6)GalNAc-R2 by a beta 6-N-acetylglucosaminyltransferase in pig gastric mucosa.
A UDP-GlcNAc:R1-beta 1-3Gal(NAc)-R2 [GlcNAc to Gal(NAc)] beta 6-N-acetylglucosaminyltransferase activity from pig gastric mucosa microsomes catalyzes the formation of GlcNAc beta 1-3(GlcNAc beta 1-6)Gal-R from GlcNAc beta 1-3Gal-R where -R is -beta 1-3GalNAc-alpha-benzyl or -beta 1-3(GlcNAc beta 1-6)GalNAc-alpha-benzyl. This enzyme is therefore involved in the synthesis of the I antigenic determinant in mucin-type oligosaccharides. The enzyme also converts Gal beta 1-3Gal beta 1-4Glc to Gal beta 1-3(GlcNAc beta 1-6)Gal beta 1-4Glc. The enzyme was stimulated by Triton X-100 at concentrations between 0 and 0.2% and was inhibited by Triton X-100 at 0.5%. There is no requirement for Mn2+ and the enzyme activity is reduced to 65% in the presence of 10 mM EDTA. Enzyme products were purified and identified by proton NMR, methylation analysis and beta-galactosidase digestion. Competition studies suggest that this pig gastric mucosal beta 6-GlcNAc-transferase activity is due to the same enzyme that converts Gal beta 1-3GalNAc-R to mucin core 2, Gal beta 1-3(GlcNAc beta 1-6)GalNAc-R, and GlcNAc beta 1-3GalNAc-R to mucin core 4, GlcNAc beta 1-3(GlcNAc beta 1-6)GalNAc-R. Substrate specificity studies indicate that the enzyme attaches GlcNAc to either Gal or GalNAc in beta (1-6) linkage, provided these residues are substituted in beta (1-3) linkage by either GlcNAc or Gal. The insertion of a GlcNAc beta 1-3 residue into Gal beta 1-3GalNAc-R to form GlcNAc beta 1-3Gal beta 1-3GalNAc-R prevents insertion of GlcNAc into GalNAc. These studies establish several novel pathways in mucin-type oligosaccharide biosynthesis.
D000818:Animals; D001667:Binding, Competitive; D003106:Colon / Q000201:enzymology; D004285:Dogs; D004492:Edetic Acid / Q000494:pharmacology; D005753:Gastric Mucosa / Q000201:enzymology*; D005964:Glucosyltransferases / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000201:enzymology; D009682:Magnetic Resonance Spectroscopy; D008745:Methylation; D008861:Microsomes / Q000201:enzymology; D009077:Mucins / Q000096:biosynthesis*; D017351:N-Acetylglucosaminyltransferases; D017830:Octoxynol; D009844:Oligosaccharides / Q000378:metabolism*; D011092:Polyethylene Glycols / Q000494:pharmacology; D013363:Submandibular Gland / Q000201:enzymology; D013379:Substrate Specificity; D013552:Swine
D009077:Mucins; D009844:Oligosaccharides; D011092:Polyethylene Glycols; D017830:Octoxynol; D004492:Edetic Acid; D005964:Glucosyltransferases; D017351:N-Acetylglucosaminyltransferases; C039548:UDP-N-acetylglucosamine-galactose(1-6)-N-acetylglucosaminyltransferase