Structural analysis of N-linked oligosaccharide of mitogenic lectin-B from the roots of pokeweed (phytolacca americana)

The structure of an asparagine (N-) linked oligosaccharide of pokeweed lectin-B (PL-B) and the amino acid sequences around two glycosylation sites were identified. The pyridylamino (PA) oligosaccharide prepared from PL-B was eluted as a single peak on both reversed-phase (RP-) HPLC and size-fractionation (SF-) HPLC, and its structure was estimated to be Manαl→6(Manαl→3)(Xylβ1→2) Manβ1→4GlcNAcβ1→4(Fucα1→3)GlcNAc by a combination of component analysis, successive exoglycosidase digestions, IS-MS analysis, and 500 MHz ¹H-NMR. Two tryptic glycopeptides were isolated from the reduced and S-pyridylethylated PL-B after gel filtration followed by RP-HPLC, indicating the presence of two glycosylation sites in PL-B. The amino acid sequences around the two glycosylation sites were determined to be Cys-Glv-Val-Asp-Phe-Gly-Asn(CHO)-Arg.