The formation of lipid-linked oligosaccharides in Madin-Darby canine kidney cells. Changes in oligosaccharide profiles induced by glucosamine.

Glucosamine inhibits the incorporation of [2-3H]mannose into lipid-linked oligosaccharides and into glycoproteins in influenza virus-infected MDCK cells. Fifty per cent inhibition of these components requires about 2 mM glucosamine. The oligosaccharide portions of the lipid-linked oligosaccharides in cells inhibited with glucosamine were compared to that of normal cells by chromatography on Bio-Gel P-4 columns. In uninhibited cells, the major oligosaccharide formed from [2-3H]mannose was the Glc3Man9GlcNAc2 species as demonstrated by the products of endoglucosaminidase H and alpha-mannosidase digestion. At low concentrations of glucosamine (approximately 2 mM) or in short term incubations (1 to 2 h), the large oligosaccharide disappeared and was replaced by a Man7GlcNAc2 species. This was also characterized by various enzymatic treatments as well as its migration rate on Bio-Gel P-4 as compared to known oligosaccharides. At still higher glucosamine concentrations or longer incubation times, the Man7GlcNAc2 species also disappeared and was replaced by a Man3GlcNAc2 species. The effect of glucosamine was reversible such that when the cells were washed free of this inhibitor, they resumed the synthesis of the Glc3Man9GlcNAc2 species and the other two oligosaccharides disappeared. These smaller oligosaccharides were not observed when glucosamine was replaced by either 5 mM galactosamine or 5 mM N-acetylglucosamine.