Abstract
The interaction of Japanese elderberry bark lectin (Sambucus sieboldiana agglutinin, SSA) with carbohydrate was investigated by 1H-NMR. When a low affinity ligand, methyl β-D-galactoside (βMeGal), was mixed with SSA, each proton signal of βMeGal was broadened. The signal of H-4 was markedly broad, while those of H-1, OCH3, and H-2 of βMeGal were rather sharp. The specific broadening of Gal H-4 was more evident when SSA was mixed with methyl-β-D-lactoside (βMeLac). Position-dependent signal broadening suggests that βMeGaI binds to SSA such that H-4 is closely involved in the contact region, but H-1, OCH3, and H-2 are far from this region. In the case of a high affinity ligand, Neu5Ac(α2-6)Gal(β1-4)Glc(=N6L), ligand signals of the SSA-N6L mixture did not change at all. But when a small amount of N6L was added to the SSA-βMeGal mixture, the broad signals of bound βMeGal became dramatically sharp. This indicates that the added N6L molecules liberated the bound βMeGal from SSA. On the other hand, the sialyllactose with the α(2-3)-linkage(=N3L) could not substitute for bound βMeGal because of its lower affinity. This demonstrates that the competitive binding experiment between two ligands is a useful technique to detect the interaction of lectins with high affinity ligands which could not be observed directly by NMR signal broadening and/or chemical shift change.
