Structural analysis of oligosaccharides ligands to a serum lectin inducing an anti-tumor activity

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Serum Mannan-binding protein (MBP), a C-type mammalian lectin specific for mannose, N-acetylglucosamine and fucose, has a potent growth inhibitory activity to a human colorectal carcinoma cell line in vivo. In this study, we have characterized the MBP-ligand oligosaccharides expressed on the human colorectal carcinoma cell line, SW 1116, which are suggested to trigger anti-tumor activity of MBP.1.FITC-labeled MBP binding to SW1116 cells was inhibited specifically by the ligand sugars, a fucose-specific lectin, and anti-Lewis A and anti-Lewis B mAbs, suggesting that MBP binds to type-I carbohydrates.2.MBP-ligand glycopeptides were isolated by a MBP affinity column from the high molecular weight glycopeptides fraction of the pronase-digest of SW 1116 cell lysates. After hydrazinolysis followed by pyridylamination of MBP-ligand glycopeptides, PA-derivatized MBP-ligand oligosaccharides were isolated again by the MBP affinity column.(1)Carbohydrate analysis indicated that MBP-ligand oligosaccharides contained high molecular size N-glycans with high galactose, N-acetylglucosamine and fucose contents.(2)Endo-β-galactosidase digestion of the MBP-ligand oligosaccharides resulted in a marked reduction of the binding activity to the MBP column together with the decrease of their molecular sizes, indicating the presence of poly N-acetyllactosamine structures. The selective removal of fucose residues from MBP-ligand oligosaccharides resulted in almost complete loss of the binding activity to the MBP and the AAL affinity columns.(3)MALDI-MSIMS and nano ESI-MS/MS analyses of the MBP-ligand oligosaccharides indicated the presence of highly fucosylated structure.MBP-ligand oligosaccharides appear to be high molecular weight poly N-acetyllactosamine-type with high fucose content.