The total content and pattern of gangliosides were determined in the unfractionated sera of 11 healthy human adults and in isolated lipoproteins. The total content of lipid-bound sialic acid was 10.5 3.2 nmol/ml serum. The ganglioside profile consisted of more than ten different components. The major ganglioside was GM3, followed by GD3, GD1,, GM2, GTlb, MG-3 (sialosyllactoneotetraosylceramide), GDlb and GQlb. Traces of four additional gangliosides could not be quantified reliably. Ganglioside patterns did not vary in sera taken from healthy adults of different age and sex. Approximately 98% of human serum gangliosides were transported by serum lipoproteins, predominantly by LDL (66%), followed by HDL (25%) and VLDL (7%). The quantitative distribution of individual gangliosides in VLDL and LDL was almost the same as that in the unfractionated serum; some differences existed with the ganglioside profile in HDL.Gangliosides are sialic-acid-containing glycosphingolipids, ubiquitous in neural and extraneural organs of deuterostomia [l]. They Svennerholm [35, 361, in parenthesis. The nomenclature for MG-3 and MG-4 is according to [17]. GM3, (I13NeuAcLacCer), (N-acetylneuraminosyl)galactosylglucosylceramide; GM2, (I13Neu-AcCgOse3Cer), N-acetylgalactosaminyl-(N-acetylneuraminosy1)galactosylglucosylceramide; G M I , (I13NeuAcGgOse4Cer), galacto-
UDP‐N‐acetylgalactosamine–GM 3acetylgalactosaminyltransferase(GM 2‐synthase) was studied in a Golgi‐rich fraction from rat liver. Activity in a cell‐free system required the presence of detergents; octyl glucoside was found to be the most effective in stimulating the enzyme. Optimal activity of GM 2‐synthase was obtained at pH 7.2, in the presence of 0.8% octyl glucoside, 10 mM Mn2+and 5 mM CDP‐choline. The latter was used to counteract the rapid sugar nucleotide hydrolysis caused by a nucleotide pyrophosphatase activity in the Golgi fraction. The apparent Kmvalue for UDP‐N‐acetylgalactosamine was 0.18 mM and Co2+and Fe2+exceeded Mn2+in activating GM 2‐synthase. Under optimal assay conditions and in the presence of added GM 3and 5 mM CDP‐choline, the specific activity of the enriched Golgi fraction was measured to be 25–30 nmol × mg protein−1× h−1; with endogenous GM 3as the sole glycolipid acceptor, V was calculated to be 9 nmol × mg protein−1× h−1.
The conditions for the quantitative determination of UDP-Gal :glucosylceramide galactosyltrai~sferase and of UDP-Gal : GM, galactosyltransferase in Golgi-enriched preparations of rat liver were optimized. Triton X-100 was the detergent routinely used as octyl glucoside acted as a galactose acceptor forming octyl lactoside. Manganese ions were required for full activity, but Co2+ and Mg2+ could substitute to some extent. The nucleotide pyrophosphatase activity of the Golgi preparations which interfered with the GL,-synthase assay was inhibited by addition of 20 rnM IMP; the latter is without appreciable effect on the rate of GL, synthesis. Apparent K, values for UDP-Gal were 130 pM and 140 pM with GI,-synthase and Gm,-synthase, respectively. That for glucosylceramide was 80 pM with GL,-synthasc; for GM, it was 10 pM with GM,-synthase. Competition experiments with variable concentrations of the lipid acceptors showed that the two synthase activities are independent catalytic entities. The specific activity of GM,-synthase exceeds that of GL,-synthase by a factor of ca. 25 under the optimized conditions used here.Gangliosides are a group of acidic glycosphingolipids mainly found in plasma membranes of neural and extraneural organs [I]. Several receptor functions were attributed to this group of lipids, but only GM, has been established so far as receptor in the case of cholera toxin [2] and, possibly, of interferon [3]. The biosynthesis of gangliosides in rat liver occurs predominantly in the Golgi apparatus [4]. Monosaccharides and sialic acids are added to the appropriate glycolipid acceptors in ordered sequence by transferases that are membrane-bound and probably organized in a multienzyine complex [S,h].Previous investigations in this laboratory showed a strong inhibition in vivo of G M I and GD,,,, biosynthesis in the liver of rats previously injected with D-galactosamine [7]. This aminosugar leads to a change in the hepatic UDP-sugar levels with lowered content of UDP-Glc and UDP-Gal and elevated content of UDP-hexosamines and UDP-N-acetylhexosamines. Ganglioside biosynthesis could, therefore, be impaired by the
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.