Three-state thermal unfolding of onconase

Onconase is a member of the ribonuclease A superfamily currently in phase IIIb clinical trials as a treatment for malign mesothelioma due to its cytotoxic activity selective against tumor-cells. In this work, we have studied the equilibrium thermal unfolding of onconase using a combination of several structural and biophysical techniques. Our results indicate that at least one significantly populated intermediate, which implies the exposure of hydrophobic surface and significant changes in the environment around Trp3, occurs during the equilibrium unfolding process of this protein. The intermediate begins to populate at about 30° below the global unfolding temperature, reaching a maximum population of nearly 60%, 10° below the global unfolding temperature.

Highlights► In the 2.0-6.0 pH range, the thermal unfolding of onconase takes place via, at least, one equilibrium intermediate. ► This intermediate implies exposure of hydrophobic surface and changes around Trp3. ► 25% of the total enthalpy change upon unfolding takes place in the native to intermediate transition. ► The stability of the intermediate is close to that of the native state.