Studies on the binding site of the galactose-specific agglutinin PA-IL from Pseudomonas aeruginosa

The binding properties of Pseudomonas aeruginosa agglutinin-I (PA-IL) withglycoproteins (gps) and polysaccharides were studied by both thebiotin/avidin-mediated microtiter plate lectin-binding assay and theinhibition of agglutinin-glycan interaction with sugar ligands. Among 36glycans tested for binding, PA-IL reacted best with two glycoproteinscontaining Galalpha1-->4Gal determinants and a human blood group ABOprecursor equivalent gp, but this lectin reacted weakly or not at all withA and H active gps or sialylated gps. Among the mammalian disaccharidestested by the inhibition assay, the human blood group PkactiveGalalpha1-->4Gal, was the best. It was 7.4-fold less active thanmelibiose (Galalpha1-->6Glc). PA-IL has a preference for thealpha-anomer in decreasing order as follows: Galalpha1-->6>Galalpha1-->4 >Galalpha1-->3. Of the monosaccharides studied,the phenylbeta derivatives of Gal were much better inhibitors than themethylbeta derivative, while only an insignificant difference was foundbetween the Galalpha anomer of methyl- and p -NO2-phenyl derivatives. Fromthese results, it can be concluded that the combining size of theagglutinin is as large as a disaccharide of the alpha-anomer of Gal atnonreducing end and most complementary to Galalpha1-->6Glc. As for thecombining site of PA-IL toward the beta-anomer, the size is assumed to beless than that of Gal; carbon-6 in the pyranose form is essential, andhydrophobic interaction is important for binding.