Onconase: An unusually stable protein

Several members of the RNase A superfamily are endowed with antitumor activ ity, showing selective cytotoxicity toward tumor cell lines. One of these i s onconase, the smallest member of the superfamily, which at present is und ergoing phase-III clinical trials as an antitumor drug. Our investigation f ocused on other interesting features of the enzyme, such as its unusually h igh denaturation temperature, its low catalytic activity, and its renal tox icity as a drug. We used differential scanning calorimetry, circular dichro ism, fluorescence measurements, and limited proteolysis to investigate the molecular determinants of the stability of onconase and of a mutant, (M23L) -ONC, which is catalytically more active than the wild-type enzyme, and ful ly active as an antitumor agent. The determination of the main thermodynami c parameters of the protein led to the conclusion that onconase is an unusu ally stable protein. This was confirmed by its resistance to proteolysis. O n the basis of this analysis and on a comparative analysis of the (M23L)-ON C variant of the protein, which is less stable and more sensitive to proteo lysis, a model was constructed in line with available data. This model supp orts a satisfactory hypothesis of the molecular basis of onconase stability and low-catalytic activity.