Thermodynamic Aspects of Fatty Acids Binding to Human Serum Albumin: A Microcalorimetric Investigation.

Abstract

Thermodynamic parameters have been evaluated for the binding interaction between human serum albumin (HSA) and unbranched fatty acids (FFA) on the basis of a flow microcalorimetric measurement at pH 7.4 and 37 degrees C by computer-fitting to single- and two-class binding models. The heat of binding increased exothermically with increasing alkyl chain length. FFA with nine or less carbons bound to only one class of binding sites (n = 2) with a binding constant (K) of 10(4) M-1. FFA with ten or more carbons bound to the first class of binding sites with high affinity K in the older of 10(5) to 10(6) M-1, and to the second class with a lower affinity and high capacity. The free energy change of the first class of binding sites (delta G1) became more negative as the chain length of FFA was increased. The enthalpy change per mol of FFA (delta H) decreased at the rate of -7.47 kJ.mol-1.CH-1(2) to a minimum at C9 and then increased due to the hydrophobicity of alkyl chains. Compensation analysis for the i th class of HSA molecule by plotting molar changes of enthalpy (delta Hmi) against entropy (delta Smi) and free energy (delta Gmi) indicates two distinct binding sites. The first class (i = 1) of the long-chain FFA on HSA is an entropy-driven reaction associated with nearly constant values of delta Hm1 (-43.0 +/- 4.8 kJ.mol-1), slightly negative values of delta Sm1 (-47.4 less than or equal to delta Sm1 less than or equal to -8.1 J.mol-1.K-1) and -delta Gm1 values, increasing with increasing alkyl chain length. The second class (i = 2) of the long-chain FFA may lie in the same region as the binding sites of the short- and medium-chain FFA with a linear relationship between delta Hmi-delta Smi.