The Asparagine-linked Oligosaccharides on Human and Porcine Plasma Vitronectins

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  • Asparagine linked Oligosaccharides on H

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The structures of asparagine-linked oligosaccharides present on plasma vitronectins of human (hVN) and porcine (pVN) are reported. Oligosac-charides were released from vitronectins by N-glycanase digestion before and after desialylation. The reducing ends of the oligosaccharides were tagged with 2-aminopyridine. Pyridylamino (PA)-derivatives of asialooligosaccharides were fractionated and analyzed by two-dimensional sugar mapping method (N. Tomiya et al. Anal. Biochem., 171, 73-90 (1988)) using reversed-phase and amide-adsorption high performance liquid chromatography (HPLC) in combination with exoglycosidase digestion. More than 70% of the total PA-asialooligosaccharides of both vitronectins were biantennary lactosamine structures. Structures of the major oligosaccharides were confirmed by 400MHz ^1H NMR spectroscopy. PA-sialooligosaccharides were fractionated by anion-exchange and reversed-phase HPLC. Ten and nine major PA-sialooligosaccharides were isolated from hVN and pVN, respectively, and after desialylation their asialo-forms were assigned to PA-asialooligosaccharides by the two-dimensional sugar mapping meth\ od. Using desialylation with Salmonella sialidase, which has a kinetic preference for sialyl α2→3 linkages, in combination with acid-desialylation, the linkage and location of sialic acids were presumed by comparing the elution position on HPLC with those of standard PA-sialooligosaccharides and standard derivatives obtained by exoglycosidase digestion. The oligosaccharides of hVN and pVN differed in extent of sialylation; 8% mono-, 70% di- and 22% tri-sialylated for hVN; 7% mono-, 89% di- and 4% tri-sialylated for pVN. Sialic acid was linked dominantly by α2→6 linkage, although α2→3 linkage was also present on both vitronectins. The most remarkable difference between hVN and pVN is the amount of fucose linked to the innermost N-acetylglucosamine by α1→6 linkage.

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