Roles of mammalian structural units, ligand cluster and polyvalency in the Abrus precatorius agglutinin and glycoprotein recognition process

Previous studies on one of the toxic type 2 ribosome inactivating proteins (RIP), Abrus precatorius agglutinin (APA), have shown that the recognition domains of APA are restricted to monomers of Galβ1-3GalNAc (T, Thomsen-Friedenreich glycotope) and Galβ1-3/4GlcNAc (blood group precursor type I/II sequences); which are essential but play a minor role in the recognition process. In this study, APA recognition factors were expanded to include ligand clusters and polyvalent glycotopes by enzyme-linked lectinosorbent binding and inhibition assays. Based on the results of molar relative potency, the essential mammalian structural units are Galβ1-3GalNAcα/β1- (Tα/Tβ) > Galα1-4Gal (E) > Galβ1-3/4GlcNAc (I/II) and avidity for tri-/di-antennary IIβ, T, E and II monomers was found to be 7.1 × 102, 4.0, 5.5, 3.7 and 2.4 times higher than monomeric Gal. Among natural polyvalent glycotopes or clusters, high-density polyvalent Tα and complex multivalent Iβ/IIβ glycotopes greatly enhanced the affinity for APA over 104 times. Based on these results, it is concluded that contribution of monomeric Tα, IIβ, Iβ, Eβ and their clusters and polyvalency play critical roles in this recognition process. The binding intensities of these factors in decreasing order are: polyvalent Tα, IIβ/Iβ and Eβ > tri-antennary IIβ ≫ monomeric Tα, Tβ, I and II > Gal ≫ GalNAc (weak). As one of type 2 RIP lectins, these recognition factors of the B chain are likely to be crucial for attachment and endocytosis. A comparison of the differential recognition factors and combining sites of APA with those of other lectins (Ricinus communis agglutinin, RCA1 and ricin) is also illustrated.