In our recent publication, we defined core aspects of the
carbohydrate specificity of domain-I of recombinant tandem-repeat-type
galectin-4 from rat gastrointestinal tract (G4-N), especially its potent interaction with the linear tetrasaccharide Galbeta1-3GlcNAcbeta1-3Galbeta1-4Glc (Ibeta1-3L). The assumed role of
galectin-4 as a microvillar raft stabilizer/organizer and as a
malignancy-associated factor in hepatocellular and gastrointestinal
carcinomas called for further refinement of its binding specificity. Thus, the effects of polyvalency of glycotopes and natural modifications of human
blood group ABH/Lewis
sugars at the terminal Galbeta1-core saccharides were thoroughly examined by the
enzyme-linked lectinosorbent and
lectin-
glycan inhibition assays. The results indicate that (a) a high-density of polyvalent Galbeta1-3/4GlcNAc (I/II), Galbeta1-3GalNAc (T) and/or GalNAcalpha1-Ser/Thr (Tn) strongly favors G4-N/glycoform binding. These
glycans were up to 2.3 x 10(6), 1.4 x 10(6), 8.8 x 10(5), and 1.4 x 10(5) more active than
Gal, GalNAc, monomeric I/II and T, respectively; (b) while lFuc is a poor inhibitor, its presence as alpha1-2 linked to terminal Galbeta1-containing
oligosaccharides, such as H active Ibeta1-3L, markedly enhances the reactivities of these
ligands; (c) when
blood group A (GalNAcalpha1-) or B (Galalpha1-) determinants are attached to terminal Galbeta1-3/4GlcNAc (or Glc)
oligosaccharides, the reactivities are also increased; (d) with lFucalpha1-3/4 linked to sub-terminal GlcNAc, the reactivities of these
haptens are reduced; and (e) short chain
Le(a)/Le(x)/Le(y) and the short chains of
sialyl Le(a)/Le(x) are poor inhibitors. These distinct binding features of G4-N establish the important concept of affinity enhancement by high density polyvalencies of glycotopes (vs. multi-antennary I/II) and by introduction of an ABH key
sugar to Galbeta1-terminated core glycotopes. The polyvalent
ligand binding properties of G4-N may help our understanding of its crucial role for cell membrane raft stability and provide salient information for the optimal design of blocking substances such as anti-tumoral glycodendrimers.