Thermodynamic studies of saccharide binding to artocarpin, a B-cell mitogen, reveals the extended nature of its interaction with mannotriose [3,6-Di-O-(α-D-mannopyranosyl)-D-mannose]

Abstract

The thermodynamics of binding of various saccharides to artocarpin, from Artocarpus integrifoliaseeds, a homotetrameric lectin (M_r 65,000) with one binding site per subunit, was determined by isothermal titration calorimetry measurements at 280 and 293 K. The binding enthalpies, ΔH_b, are the same at both temperatures, and the values range from −10.94 to −47.11 kJ mol⁻¹. The affinities of artocarpin as obtained from isothermal titration calorimetry are in reasonable agreement with the results obtained by enzyme-linked lectin absorbent essay, which is based on the minimum amount of ligand required to inhibit horseradish peroxidase binding to artocarpin in enzyme-linked lectin absorbent essay (Misquith, S., Rani, P. G., and Surolia, A. (1994) J. Biol. Chem. 269, 30393-30401). The interactions are mainly enthalpically driven and exhibit enthalpy-entropy compensation. The order of binding affinity of artocarpin is as follows: mannotriose > Manα 3Man > GlcNAc₂Man₃ > MeαMan > Man > Manα6Man > Manα2Man > MeαGlc > Glc,i.e. 7 > 4 > 2 > 1.4 > 1 > 0.4 > 0.3 > 0.24 > 0.11. The ΔH for the interaction of Manα3Man, Manα6Man, and MeαMan are similar and 20 kJ mol⁻¹ lower than that of mannotriose. This indicates that, while Manα3Man and Manα6Man interact with the lectin exclusively through their nonreducing end monosaccharide with the subsites specific for the α1,3 and α1,6 arms, the mannotriose interacts with the lectin simultaneously through all three of its mannopyranosyl residues. This study thus underscores the distinction in the recognition of this common oligosaccharide motif in comparison with that displayed by other lectins with related specificity.