Structural stabilization of bovine β-Lactoglobulin in presence of polyhydric alcohols

Abstract

β-Lactoglobulin (β-Lg) is the major protein present in bovine milk whey. Addition of hydrogen-bonded cosolvents to proteins is known to modify the thermodynamic properties of proteins. Preferential interaction parameters of β-Lg were determined in various concentrations of cosolvents like sorbitol, glycerol and sucrose using the precision densitymetry. The apparent partial specific volumes determined at 20°C under both isomolal and isopotential conditions in 0.02 M phosphate buffer (pH 7.9) were 0.743±0.001 and 0.744±0.001, respectively. From the partial specific volume data with cosolvents the preferential interaction parameter and other thermodynamic parameters were calculated at different cosolvent concentrations. The values increased with solvent concentration up to 40% and reached a maximum of 0.190±0.02 and 0.140±0.02 in sucrose and sorbitol, respectively. In glycerol, the value increased up to 0.213±0.03 in 20% glycerol and then decreased with the increase in the cosolvent concentration. There were no changes in secondary structure of the protein as reflected by far UV-CD spectra. The fluorescence spectra showed changes in the fluorescence intensity due to perturbations in the tryptophan moiety without changes in the emission wavelength. The above data was further supported by the degree of hydrolysis in the presence of cosolvents with the enzyme ⍺-chymotrypsin. The degree of hydrolysis reduced to 12% from the control value of 18% in the presence of sorbitol and glycerol and increased by 3% in the presence of sucrose. All the cosolvents also enhanced thermal stability to various extents.