Counteraction ability of TMAO toward different denaturing agents

Differential scanning calorimetry measurements performed on RNase A in aqueous binary solutions containing different concentrations of urea, tetramethylurea, guanidinium chloride, and guanidinium thiocyanate, and in aqueous ternary solutions, containing the same denaturants plus 1 M trimethylamine N-oxide, TMAO, demonstrate that the latter has a general counteracting ability at pH 7.0, but not at pH 4.0. Experimental data rule out the idea that counteraction originates from direct interactions between TMAO molecules and denaturing agents. A rationalization is provided on the basis of a theoretical approach grounded on the solvent-excluded volume effect, whose magnitude depends on the density of aqueous solutions.