Carbohydrate structures of soluble human L-selectin recombinantly expressed in baby-hamster kidney cells

A soluble form of L-selectin was recombinantly produced, which might be an effective therapeutic agent in inflammatory disorders, acting as an inhibit or for leucocyte endothelium adhesion, In the present study the oligosaccha ride structures of soluble human L-selectin, recombinantly expressed in bab y-hamster kidney cells, were determined. The N-linked glycans were enzymica lly released and fluorescently labelled with 2-aminobenzamide. Sialylation of the N-glycans was analysed by anion-exchange chromatography followed by rechromatography of the resulting fractions on amino-phase HPLC after relea se of the sialic acid residues. Desialylated oligosaccharides were separate d using two-dimensional HPLC and characterized by digestion with exoglycosi dases and MS. More than 30 oligosaccharide structures representing at least 95 % of the overall glycosylation of this protein were determined. The res ults revealed that recombinant soluble human L-selectin carries bi-, tri- a nd tetra-antennary sugar chains, which are fucosylated on the innermost res idue of N-acetylglucosamine, The number of sialic acid residues linked to t hese glycans ranges from 9 (neutral glycans) to 4 (tetrasialylated oligosac charides). The sialic acid is found exclusively in the alpha 2-3 linkage to galactose. In addition to the main glycans, different minor structures con taining terminal N-acetylgalactosamine, or the H (O) blood-group determinan t were also identified. O-Glycosylation of mucin-type sugar chains was not detected in recombinant soluble human L-selectin.