Lectin affinity high-performance liquid chromatography: Interactions of N-glycanase-released oligosaccharides with leukoagglutinating phytohemagglutinin, concanavalin A, Datura stramonium agglutinin, and Vicia villosa agglutinin (original) (raw)

1987, Analytical Biochemistry

The structural determinants required for interaction of oligosaccharides with Ricinus communis agglutinin I (RCA]) and Ricinus communis agglutinin I1 (RCAII) have been studied by lectin affinity high-performance liquid chromatography (HPLC). Homogeneous oligosaccharides of known structure, purified following release from Asn with N-glycanase and reduction with NaBH4, were tested for their ability to interact with columns of silica-bound RCAl and RCAII. The characteristic elution position obtained for each oligosaccharide was reproducible and correlated with specific structural features. RCAI binds oligosaccharides bearing terminal /31,4-linked Gal but not those containing terminal j31,4-linked GalNAc. In contrast, RCAII binds structures with either terminal B1,4-linked Gal or B1,4-linked GalNAc. Both lectins display a greater affinity for structures with terminal /31,4-rather than @1,3-linked Gal, although RCAII interacts more strongly than RCAl with oligosaccharides containing terminal @1,3-linked Gal. Whereas terminal a2,6linked sialic acid partially inhibits oligosaccharide-RCAI interaction, terminal a2,3-linked sialic acid abolishes interaction with the lectin. In contrast, a2,3-and a2,g-linked sialic acid equally inhibit but do not abolish oligosaccharide interaction with RCAII. RCAI and RCAIl discriminate between N-acetyllactosamine-type branches arising from different core Man residues of dibranched complex-type oligosaccharides; RCAI has a preference for the branch attached to the al,3-linked core Man and RCAIl has a preference for the branch attached to the al,g-linked core Man. RCAIl but not RCAI interacts with certain di-and tribranched oligosaccharides devoid of either Gal or GalNAc but bearing terminal GlcNAc, indicating an important role for GlcNAc in RCAII interaction. These findings suggest that N-acetyllactosamine is the primary feature required for oligosaccharide recognition by both RCAI and RCAII but that lectin interaction is strongly modulated by other structural features. Thus, the oligosaccharide specificities of RCAI and RCAII are distinct, depending on many different structural features including terminal sugar moieties, peripheral branching pattern, and sugar linkages. Grant R37-CA21923 and by the Monsanto Company/Washington * This investigation was supported by National Cancer Institute University Biomedical Research Grant. The costs of publication of this article were defrayed in part by the payment of page charges.