Locating the binding sites of citric acid and gallic acid on milk β-lactoglobulin

Abstract

β-Lactoglobulin (β-LG) is a member of lipocalin superfamily of transporters for small hydrophobic and hydrophilic molecules. We located the binding sites of citric acid and gallic acid on β-lactoglobulin (β-LG) in aqueous solution at physiological conditions, using spectroscopic methods, thermodynamic analysis and molecular modeling. Thermodynamic parameters ΔH0 −9.5 to −6.9 (kJ mol−1), ΔS0 23.9 to 13.6 (J mol−1K−1) and ΔG0 −14.5 to −13.6 (kJ mol−1) showed that acid binds protein via ionic contacts with gallic acid forming stronger protein conjugates consistent with theoretical modeling. Different amino acids are involved in gallic acid and citric acid complexation. Protein conformation was altered with reduction of β-sheet from 58% (free protein) to 46–43% and a major increase in α-helix from 11% (free protein) to 29–23% and random coil structure in the acid-protein, indicating a partial protein destabilization. Communicated by Ramaswamy H. Sarma