ABSTRACT
Urine from Sd(a+) individuals was found to contain a beta-N-acetylgalactosaminyltransferase that transfers N-acetylgalactosamine (GalNAc) from UDP-GalNAc to 3'-sialyllactose and glycoproteins carrying the terminal NeuAc alpha-3Gal beta group. This enzyme has been purified 174-fold by affinity chromatography on Blue Sepharose and DEAE-Sephacel chromatography in a yield of 33%. Neither endogenous incorporation nor sugar nucleotide degrading enzymes were found in the purified preparation. The transferase had a pH optimum of pH 7.5 and a requirement for Mn2+ but not for detergents. The Km for UDP-GalNAc was 66 X 10(-6) M, using fetuin as an acceptor. Like beta-GalNAc-transferase from other sources the urinary enzyme had a strict requirement for sialylated acceptors. On the basis of enzymatic and chemical treatment of the product obtained by the transfer of [3H]GalNAc to 3'-sialyllactose, we propose that the enzyme attaches GalNAc in beta-anomeric configuration to O-4 of the galactose residue that is substituted at O-3 by sialic acid. A preparation of Tamm-Horsfall glycoprotein from a Sd(a-) donor lacking beta-GalNAc was found to be the best acceptor among the glycoproteins tested. Studies on the transferase activity toward fetuin, human chorionic gonadotropin, and glycophorin A indicated that the enzyme preferentially adds the sugar to the sialylated terminal end of N-linked oligosaccharides. Unlike the beta-GalNAc-transferase bound to human kidney microsomes (F. Piller et al. (1986) Carbohydr. Res. 149, 171-184) the urinary transferase is able to transfer beta-GalNAc to the NeuAc alpha-3Gal beta-3(NeuAc alpha-6)GalNAc chains bound to the native glycophorin.
Subject(s)
Blood Group Antigens , Galactosyltransferases/urine , N-Acetylgalactosaminyltransferases , Chromatography, Affinity , Chromatography, Gel , Glycoproteins/metabolism , Humans , Oligosaccharides/metabolism , Substrate SpecificityABSTRACT
alpha-Galactosyltransferase which participates in the biosynthesis of blood group B substance was found in urine from group B and AB healthy persons of both secretors and non-secretors. The activity of alpha-galactosyltransferase in the urine of a healthy variant Bm person was lower than that found in a normal group B person. This enzyme in urine of A1Bm persons must be much lower than that in normal AB persons. Its activity was not detected by the method of group O to group B transformation or erythrocytes.
Subject(s)
ABO Blood-Group System/genetics , Galactosyltransferases/urine , Galactosyltransferases/genetics , Genetic Variation , HumansABSTRACT
By chromatographic method we have shown the existence of a complex system for galactose transfer from UDP-galactose and for nucleotide hydrolysis in urines from Balb/c YC8 and normal Balb/c mice. By action of sera from normal and ascitic mice as source of enzyme, we have been able to detect transfer for galactose in urines from ascitic mice and an important inhibitory effect of the nucleotide sugar hydrolysis by the sera with urines from normal mice.
