Evidence that Water Can Reduce the Kinetic Stability of Protein-Hydrophobic Ligand Interactions

The first quantitative comparison of the thermal dissociation rate constants measured for protein-ligand complexes in their hydrated and dehydrated states is described. Rate constants, measured using surface plasmon resonance spectroscopy, are reported for the dissociation of the 1:1 complexes of bovine beta-lactoglobulin (Lg) with the fatty acids (FA), palmitic acid (PA), and stearic acid (SA), in aqueous solution at pH 8 and at temperatures ranging from 5 to 45 degrees C. The rate constants are compared to values determined from time-resolved blackbody infrared radiative dissociation measurements for the gaseous deprotonated (Lg+FA)(n-) ions, where n = 6 and 7, at temperatures ranging from 25 to 66 degrees C. Notably, the hydrated (Lg+PA) complex is kinetically less stable than the corresponding gas phase (Lg+PA)(n-) ions at all temperatures investigated; the hydrated (Lg+SA) complex is kinetically less stable than the gaseous (Lg+SA)(n-) ions at temperatures <45 degrees C. The greater kinetic stability of the gaseous (Lg+FA)(n-) ions originates from significantly larger, by 11-12 kcal mol(-1), E-a values. It is proposed that the differences in the dissociation E-a values measured in solution and the gas phase reflect the differential hydration of the reactant and the dissociative transition state.