Solubilization and properties of mannose and N-acetylglucosamine transferases involved in formation of polyprenyl-sugar intermediates.
A particulate fraction from porcine aorta catalyzed the incorporation of N-acetylglucosamine (GlcNAc) from UDP-[3H]GlcNAc into both GlcNAc-pyrophosphorylpolyprenol and GlcNAc-GlcNAc-pyrophosphorylpolyprenol. This transfer utilized endogenous lipid and required a divalent cation. Mn2+ was the best metal ion and was optimum at 2.3 mM. This same particulate fraction was previously shown to transfer mannose from GDP-[14C]mannose to endogenous lipid to form mannosylphosphorylpolyprenol (Chambers, J., and Elbein, A.D. (1975) J. Biol. Chem. 250, 6904-6915). Both the GlcNAc activities and the mannose activity were solubilized by treatment of the particulate fraction with the detergent Nonidet P-40. The enzymes were partially purified by chromatography on DEAE-cellulose and on Sephadex G-200. These soluble enzymes required the addition of acceptor lipid for activity. An acidic lipid fraction, isolated from pig liver and having the properties of dolichyl phosphate, was active with either the GlcNAc or the mannose transferase. Chemically synthesized dolichyl phosphate was also active with either of these enzymes. The products formed from either GlcNAc or mannose by the soluble transferases were similar to those formed by the particulate enzyme. Thus the major product formed from UDP-[3H]GlcNAc was GlcNAc-pyrophosphoryldolichol with small amounts of the disaccharide-lipid while the product formed from GDP-[14C]mannose was mannosylphosphoryldolichol.
D000818:Animals; D001011:Aorta / Q000201:enzymology*; D004286:Dolichols / Q000031:analogs & derivatives / Q000096:biosynthesis; D006600:Hexosephosphates / Q000096:biosynthesis; D006602:Hexosyltransferases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008055:Lipids / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008364:Mannosyltransferases / Q000378:metabolism; D008861:Microsomes / Q000201:enzymology; D012995:Solubility; D013347:Subcellular Fractions / Q000201:enzymology; D013403:Sugar Phosphates / Q000378:metabolism*; D013552:Swine; D013729:Terpenes / Q000378:metabolism*; D014537:Uridine Diphosphate N-Acetylglucosamine
D004286:Dolichols; D006600:Hexosephosphates; D008055:Lipids; D013403:Sugar Phosphates; D013729:Terpenes; D008345:Manganese; D014537:Uridine Diphosphate N-Acetylglucosamine; D006602:Hexosyltransferases; D008364:Mannosyltransferases