IDENTIFICATION OF THE SIALOSYL-SIALOSYL LINKAGE IN BIOSYNTHESIZED GANGLIOSIDES

Article abstract of DOI:10.1016/S0008-6215(00)80747-8

A technique was developed to examine the sialosyl-sialosyl linkage in the minute quantities of complex gangliosides biosynthesized in vitro.The disialoganglioside, G_D3, and the trisialoganglioside, G_T1a, were biosynthesized as secondary products by the reaction of CMP-(4-¹⁴C)sialic acid with the glycolipid substrates, lactosyl ceramide and G_M1 ganglioside, respectively.Following periodate oxidation-borohydride reduction, the intact sialic acid residue, and the susceptible sialic acid residue converted into an N-acetylheptulosaminic acid residue, were released by hydrolysis, esterified, separated, and analyzed for radioactivity.Analysis of the biosynthesized G_D3 and G_T1a gangliosides indicated radioactivity in both the susceptible and the nonsusceptible sialic acid residues, demonstrating that a sialosyl-(2->8)-sialosyl linkage had been synthesized.When the primary products (G_M3 and G_D1a, respectively) of the synthesis just described were examined, radioactivity was detected, as expected, only in the susceptible sialic acid residue.Because the N-acetylneuraminic acid and N-acetylheptulosaminic acid derivatives formed in the reaction were identified by chromatography, the standards used for comparison were verified by g.l.c.-m.s.