Elucidation of binding specificity of Jacalin toward O-glycosylated peptides: quantitative analysis by frontal affinity chromatography.

Jacalin, a lectin from the jackfruit Artocarpus integrifolia, has been known as a valuable tool for specific capturing of O-glycoproteins such as mucins and IgA1. Though its sugar-binding preference for T/Tn-antigens is well established, its detailed specificity has not been elucidated. In this study, we prepared a series of mucin-type glycopeptides using human glycosyltransferases, that is, ST6GalNAc1, Core1Gal-T1 and -T2, beta 3Gn-T6, and Core2GnT1, and investigated their binding to immobilized Jacalin by frontal affinity chromatography ( FAC). As a result, consistent with the previous observation, Jacalin showed high affinity for T-antigen ( Core1) and Tn-antigen ( alpha N-acetylgalactosamine)attached peptides. Furthermore, we here show as novel findings that ( 1) Jacalin also showed significant affinity for Core3 and sialyl-T ( ST)-attached peptides, but ( 2) Jacalin could not bind to Core2, Core6, and sialyl-Tn ( STn)-attached peptides. The results were also confirmed by FAC using p-nitrophenyl ( pNP)-derivatized saccharides. In conclusion, Jacalin binds to a GalNAc alpha 1-peptide, in which C6-OH of alpha GalNAc is free (i.e., Core1, Tn, Core3, and ST), whereas it cannot recognize a GalNAc alpha 1-peptide with a substitution at the C6 position ( i.e., Core2, Core6, and STn). These findings provide useful information when applying jacalin for functional analysis of mucin-type glycoproteins and glycopeptides.