Biosynthesis of oligosaccharide-lipid in Streptococcus sanguis

An oligosaccharide-lipid containing N-acetyl d-glucosamine (GlcNAc), l-rhamnose, and d-glucose was synthesized when the particulate enzyme from Streptococcus sanguis was incubated with UDP-GlcNAc, TDP-rhamnose, and UDP-glucose. The incorporation of d-glucose into the lipid was dependent on the preincorporation of l-rhamnose, which in turn was dependent on that of GlcNAc. This indicates that the order of sugar incorporation is GlcNAc, l-rhamnose, and d-glucose. The synthesis of GlcNAc-lipid was stimulated twofold by ATP and was inhibited strongly by UDP and slightly by UMP, CDP, and TDP, but not by all other nucleoside diphosphates and nucleoside monophosphates tested. A [gamma-(32)P]ATP labeling experiment indicated that some acceptor lipid was present in nonphosphorylated form. The acid and alkaline stabilities of the GlcNAc-lipid were similar to those of glycosyl undecaprenylphosphate, and the thin-layer chromatographic mobility of the lipid was slightly faster than that of the mannosylphosphorylundecaprenol. The molar ratio of phosphate to GlcNAc in purified GlcNAc-lipid was found to be 0.96:1. These results suggested that the GlcNAc was attached to the lipid moiety, presumably undecaprenol, by phosphodiester bonds. The incorporation of l-rhamnose into the lipid was inhibited by UDP and UMP, respectively, in a manner similar to the incorporation of GlcNAc. This suggested that the oligosaccharide was also linked to the lipid moiety by phosphodiester bonds.