A thermodynamic study of the trp repressor-operator interaction.

We have measured the heats of formation of the trp repressor/operator complex by direct titration calorimetry over the temperature range 10 degrees C to 40 degrees C. A primary strong mode of binding displays the characteristic large negative heat capacity change observed by other methods in the formation of specific protein/DNA complexes. Unlike most such reactions, however, the formation of the trp repressor/operator complex is enthalpically driven throughout the physiological temperature range. After saturation of this principal mode, we also detected a secondary weaker binding mode, which we ascribe to a now well documented interaction called "half-site" binding. Although weak, this mode also exhibits an unusually large negative heat capacity change. Since the interface of the proposed secondary half-site binding mode has the same complementary stereochemistry as the primary one (due to internal symmetry), we correlate the negative heat capacity change with the formation of a stereospecific interface and not with high affinity. As in similar cases, the empirical correlation between buried non-polar surfaces and reduction of heat capacity does not account for the large negative delta Cp, nor do crystal structures reveal any further reduction in solvent excluded surfaces within the reactants upon complex formation. We attribute the "unaccounted for" decrement in the heat capacity of the complex to the stereospecific restriction of the hydrated polar elements that form the specific interface. We suggest that the "tightening of soft internal modes" at and near the polar interface of the complex is more important than previously recognized because previous considerations did not take into account the highly hydrated nature of these polar elements and the concomitant reduction in the degrees of freedom of the water structure.