Thermodynamics of monosaccharide binding to concanavalin A, pea (Pisum sativum) lectin, and lentil (Lens culinaris) lectin

Abstract

Titration calorimetry measurements of the binding of methyl α-D-mannopyranoside (MeαMan), D-mannopyranoside (Man), methyl α-D-glucopyranoside (MeαGlu), and D-glucopyranoside (Glu) to concanavalin A (Con A), pea lectin, and lentil lectin were performed at 281 and 292 K in 0.01 M dimethylglutaric acid-NaOH buffer (pH 6.9) containing 0.15 M NaCl and Mn⁺² and Ca⁺² ions. The site binding enthalpies, ΔH, are the same at both temperatures and range from −28.4±0.9 (MeαMan) to −16.6±0.5 kJ mol⁻¹ (Glu) for Con A, from −26.2±1.1 (MeαMan) to −12.8±0.4 kJ mol⁻¹ (MeαGlu) for pea lectin, and from −16.6±0.7 (MeαMan) to −8.0±0.2 kJ mol⁻¹ (MeαGlu) for lentil lectin. The site binding constants range from 17±1×10³M⁻¹ (MeαMan to Con A at 281.2 K) to 230±20 M⁻¹ (Glu to lentil lectin at 292.6 K) and exhibit high specificity for Con A where they are in the MeαMan:Man:MeαGlu:Glu ratio of 21:4:5:1, while the corresponding ratio is 5:2:1.5:1 for pea lectin and 4:2:2:1 for lentil lectin. The higher specificity for Con A indicates more interactions between the amino acid residues at the binding site and the carbohydrate ligand than for the pea and lentil lectin-carbohydrate complexes. The carbohydrate-lectin binding results exhibit enthalpy-entropy compensation in that ΔH_b (kJ mol⁻¹)=−1.67±0.06×10⁴+(1.30±0.12)T(K) ΔS_b (J mol⁻¹K⁻¹). Differential scanning calorimetry measurements on the thermal denaturation of the lectins and their carbohydrate complexes show that the Con A tetramer dissociates into monomers, while the pea and lentil lectin dimers dissociate into two submonomer fragments. At the denaturation temperature, one carbohydrate binds to each monomer of Con A and the pea and lentil lectins. Complexation with the carbohydrate increases the denaturation temperature of the lectin and the magnitude of the increases yield binding constants in agreement with the determinations from titration calorimetry.