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, beta3Gn-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 GalNAcalpha1-peptide, in which C6-OH of alphaGalNAc is free (i.e., Core1, Tn, Core3, and ST), whereas it cannot recognize a GalNAcalpha1-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.
Tachibana
Kouichi
K
Glycogene Function Team, National Institute of Advanced Industrial Science and Technology (AIST), Central-2, 1-1-1 Umezono, Tsukuba, Ibaraki 305-8568, Japan.
Nakamura
Sachiko
S
Wang
Han
H
Iwasaki
Hiroko
H
Tachibana
Kahori
K
Maebara
Kanako
K
Cheng
Lamei
L
Hirabayashi
J
J
Narimatsu
H
H
D015295:Antigens, Tumor-Associated, Carbohydrate / Q000737:chemistry* / Q000378:metabolism; D002846:Chromatography, Affinity; D005964:Glucosyltransferases / Q000737:chemistry*; D006801:Humans; D009077:Mucins / Q000737:chemistry* / Q000378:metabolism; D009842:Oligopeptides / Q000737:chemistry* / Q000378:metabolism; D037121:Plant Lectins / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D013379:Substrate Specificity
D015295:Antigens, Tumor-Associated, Carbohydrate; D009077:Mucins; D009842:Oligopeptides; D037121:Plant Lectins; C040286:Tn antigen; C043696:jacalin; D005964:Glucosyltransferases