Chemical Structures of Two Subunits, A-subunit and B-subunit, of Galactose-specific Isolectins from Erythrina Variegata Seeds

We previously isolated galactose-specific isolectins, EVLI, EVLII, and EVLIII, from the Erythrina variegata seeds [J. Chromatogr. 597, 207-211 (1992)]. The amino acid sequences of the two subunits, A- and B-subunits, of which the isolectins are composed, were determined. A comparison of the amino acid sequences of tryptic peptides from the two subunits revealed seven amino acid substitutions. Among them, Asn46, to which the oligosaccharide chain is linked in the A-subunit, is replaced by Asp46 in the B-subunit, causing the B-subunit to lack one glycosylation site. The N-linked oligosaccharides of these subunits were also analyzed. The N-linked oligosaccharides were first liberated by hydrazinolysis. After N-acetylation, the reducing ends of the oligosaccharides were coupled with 2-aminopyridine, and then the pyridylamino (PA-) derivatives were purified by gel filtration and HPLC on an ODS-silica column. One major sugar chain, accounting for more than 98% of the total, was purified from both species. The structure of this major sugar chain was established to be Man alpha 6(Man alpha 3)(Xyl beta 2)Man beta 4GlcNAc beta 4(Fuc alpha 3)GlcNAc. This finding that there is no structural difference of the sugar chains linked to the two subunits of E. variegata lectins, together with the results of amino acid sequence comparisons, indicates that the difference in molecular mass of these two subunits results almost wholly from the difference in the number of oligosaccharides linked to them.