Protein-ligand interaction. A calorimetric study of the interaction of oligosaccharides and hen ovalbumin glycopeptides with concanavalin A

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Abstract
A calorimetric study is reported concerning the interaction between concanavalin A (Con A) and some oligosaccharides and glycopeptides hydrolyzed from hen ovalbumin. The measurements were carried out in acetate buffer, pH 4.5, where, by far, the prevailing form of the protein is the dimeric one [Kalb, A. J., and Lustig, A. (1968) Biochim. Biophys. Acta 168, 366; Dani, M., Manca, F., and Rialdi, G. (1981) Biochim. Biophys. Acta 667, 108]. The calorimetric technique allows the direct determination of the binding enthalpy, the evaluation of the apparent association constant KB'', and then the evaluation of the apparent free energy entropy. Three groups of data have been collected in the present study. The first one concerns the interaction between concanavalin A and some mono- and disaccharides [methyl .alpha.-glucopyranoside (.alpha.MGlup), methyl .alpha.-mannopyranoside (.alpha. MManp), D-maltose, D-trehalose, and D-cellobiose]. The analysis of the data indicates that in these cases there are small favorable entropic and enthalpic contributions to the affinity. The stoichiometry of the reaction is 2 mol of ligand/mol of Con A dimer, the sites resulting being equivalent and noninteracting. Melezitose, the only trisaccharide studied, shows a different behavior: its affinity for Con A is higher as compared to the other oligosaccharides containing .alpha.-glucosyl residues and closer to that of methyl .alpha.-mannopyranoside. However, the stoichiometry is different, namely, 1 mol of ligand/dimer of Con A. The third group of ligands studied is formed by three derivatives of some branched mannans. The values of the corresponding KB'' and the other thermodynamic parameters indicate the formation of strong complexes, with a stoichiometry of 1 mol of ligand/dimer of Con A. The analysis of the thermodynamic parameters obtained allows formulation of a new hypothesis on the mechanism of the association process of glycoconjugates to this lectin; namely, the surface of Con A can adapt to a "polyfunctional" branched saccharide or glycoconjugate having at least three sugar rings with two terminal .alpha.-linked manno- or glucopyranoside rings. In the absence of this structure of the ligand, only the part of the active site operates, which can bind .alpha.-manno- and .alpha.-glucopyranosyl derivatives. The main basis of our hypothesis, which excludes the presence of an extended site, is based on the anticooperative effect promoted on the site of the second subunit of Con A dimer by the adsorption of the ligand on the site of the first subunit.