In this study the pH dependence of the thermodynamic stability of tendamistat is analyzed. This small globular protein of 74 residues shows a very marked dependence of thermal stability on pH: the denaturation temperature increases from 68.9 degrees C at pH 2.0 to 93.2 degrees C at pH 5.0, and then decreases to 77.8 degrees C at pH 8.0. Analysis of the data indicates that the binding of two protons is coupled to the thermal unfolding at pH values below 4.0, whereas one proton is released by the protein at pH values above 5.0. By linking the proton binding to the conformational unfolding equilibrium, a thermodynamic model, which is able to describe the dependence upon the solution pH of the denaturation Gibbs energy change for tendamistat, is developed.