Intracellular Localization of Liver Sugar Nucleotide Glycoprotein Glycosyltransferases in a Golgi-rich Fraction

Abstract

Abstract The following trisaccharide unit terminates the oligosaccharide chains of several plasma glycoproteins: sialic acid → galactose → N-acetylglucosamine → oligosaccharide → protein. Previous studies showed that this trisaccharide unit was synthesized by the sequential action of three glycosyl-transferases isolated from goat colostrum; each glycosyltransferase catalyzed the transfer of a monosaccharide residue (N-acetylglucosamine, galactose, or sialic acid) from its nucleotide derivative to the appropriate glycoprotein acceptor. The present studies are concerned with the intracellular location of these glycosyltransferases in rat liver. Kinetic studies were first performed to establish optimum conditions for determining quantitatively the particle-bound rat liver glycosyltransferases; these enzymes exhibited similar properties to the soluble, partially purified glycosyltransferases previously obtained from goat colostrum. The subcellular localization of the enzymes was then investigated by two techniques, differential centrifugation and discontinuous sucrose density gradient centrifugation. In these studies, the following were used as markers for various subcellular organelles: DNA, RNA, glucose 6-phosphatase, NADPH-cytochrome c reductase, acid phosphatase, 5'-nucleotidase, and glutamic dehydrogenase. The results indicated that the glycosyltransferases were all located in the same membranous subcellular component and that this component was different from organelles containing the above marker substances; the active subcellular particle was characterized by electron microscopy as being rich in Golgi apparatus. The apparent location of the glycosyltransferases in the Golgi apparatus suggests that these enzymes may be involved in terminating the synthesis of plasma glycoproteins by the liver during secretion, and may possibly be required for secretion of these proteins.