Lipid-mediated glycosylation in human liver. Characterization of the enzymatic transfer of N-acetylglucosamine from UDP-N-acetylglucosamine and mannose from GDP-mannose to dolichyl phosphate.
The enzymatic transfer of GlcNAc from UDP-GlcNAc and Man from GDP-Man to Dol-P has been characterized in human liver preparations. The presence of low concentrations of detergent, divalent cation and exogenous Dol-P are required for both enzymatic activities. The pH optimum of both reactions is broad with maximal activity near pH 7.8. The majority of N-acetylglucosaminyltransferase (90%) and mannosyltransferase (85%) activities is particulate but approximately 90% of both activities can be released into supernatant fluids by using Triton X-100 in the homogenizing buffer. The supernatant fluid enzymes have properties similar to those of the particulate enzymes although their activities are considerably less stable. Preliminary characterization of the enzymatic reaction products gave the following evidence for formation of GlcNAc and Man derivatives of Dol-P: (1) radiolabelled products are soluble in organic solvents; (2) for each reaction no detectable product is found without addition of exogenous Dol-P and increasing amounts of product are found with increasing amounts of this lipid; (3) acid and base hydrolysis of the glycolipid product (from the N-acetylglucosaminyltransferase reaction) result in radioactive, water-soluble compounds which comigrate with authentic GlcNAc and GlcNAc-1-P, respectively; (4) acid and base hydrolysis of the glycolipid product (from the mannosyltransferase reaction) result in radioactive, water-soluble compounds which comigrate with authentic Man and Man-1-P, respectively.
D004288:Dolichol Phosphates / Q000378:metabolism; D005964:Glucosyltransferases / Q000378:metabolism; D006023:Glycoproteins / Q000096:biosynthesis*; D006155:Guanosine Diphosphate Mannose / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D050356:Lipid Metabolism; D008099:Liver / Q000378:metabolism*; D008364:Mannosyltransferases / Q000378:metabolism; D017351:N-Acetylglucosaminyltransferases; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism
D004288:Dolichol Phosphates; D006023:Glycoproteins; C028306:dolichol monophosphate; D006155:Guanosine Diphosphate Mannose; D014537:Uridine Diphosphate N-Acetylglucosamine; D005964:Glucosyltransferases; D008364:Mannosyltransferases; D017351:N-Acetylglucosaminyltransferases; C037649:N-acetyllactosaminide beta-1,6-N-acetylglucosaminyltransferase