Onconase: an unusually stable protein.


Abstract

Several members of the RNase A superfamily are endowed with antitumor activity, showing selective cytotoxicity toward tumor cell lines. One of these is onconase, the smallest member of the superfamily, which at present is undergoing phase-III clinical trials as an antitumor drug. Our investigation focused on other interesting features of the enzyme, such as its unusually high denaturation temperature, its low catalytic activity, and its renal toxicity as a drug. We used differential scanning calorimetry, circular dichroism, 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 fully active as an antitumor agent. The determination of the main thermodynamic parameters of the protein led to the conclusion that onconase is an unusually stable protein. This was confirmed by its resistance to proteolysis. On the basis of this analysis and on a comparative analysis of the (M23L)-ONC variant of the protein, which is less stable and more sensitive to proteolysis, a model was constructed in line with available data. This model supports a satisfactory hypothesis of the molecular basis of onconase stability and low-catalytic activity. Study holds ProTherm entries: 8561, 8562, 8563, 8564, 8565, 8566, 8567, 8568, 8569, 8570, 8571, 8572, 8573, 8574, 8575, 8576, 8577, 8578, 8579, 8580, 8581, 8582, 8583, 8584, 8585, 8586, 8587, 8588, 8589, 8590, 8591, 8592, 8593, 8594, 8595, 8596, 8597, 8598, 8599, 8600 Extra Details: antitumor activity; catalytic activity; renal toxicity; proteolysis

Submission Details

ID: McNaD98d3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:36 p.m.

Version: 1

Publication Details
Notomista E;Catanzano F;Graziano G;Dal Piaz F;Barone G;D'Alessio G;Di Donato A,Biochemistry (2000) Onconase: an unusually stable protein. PMID:10913282
Additional Information

Number of data points 122
Proteins Protein P-30 ; Protein P-30
Unique complexes 2
Assays/Quantities/Protocols Experimental Assay: Cm ; Experimental Assay: m ; Experimental Assay: dG_H2O ; Experimental Assay: Cm ; Experimental Assay: m ; Experimental Assay: dG_H2O ; Experimental Assay: dG pH:8.0 ; Experimental Assay: dG pH:7.0 ; Experimental Assay: dG pH:6.0 ; Experimental Assay: dG pH:5.0 ; Experimental Assay: dG pH:4.0 ; Experimental Assay: dG pH:3.5 ; Experimental Assay: dG pH:3.0 ; Experimental Assay: dG pH:2.5 ; Experimental Assay: dG pH:2.0 ; Experimental Assay: dCp pH:8.0 ; Experimental Assay: dHcal pH:8.0 ; Experimental Assay: Tm pH:8.0 ; Experimental Assay: dHvH pH:8.0 ; Experimental Assay: dCp pH:7.0 ; Experimental Assay: dHcal pH:7.0 ; Experimental Assay: Tm pH:7.0 ; Experimental Assay: dHvH pH:7.0 ; Experimental Assay: dCp pH:6.0 ; Experimental Assay: dHcal pH:6.0 ; Experimental Assay: Tm pH:6.0 ; Experimental Assay: dHvH pH:6.0 ; Experimental Assay: dCp pH:5.0 ; Experimental Assay: dHcal pH:5.0 ; Experimental Assay: Tm pH:5.0 ; Experimental Assay: dHvH pH:5.0 ; Experimental Assay: dCp pH:4.0 ; Experimental Assay: dHcal pH:4.0 ; Experimental Assay: Tm pH:4.0 ; Experimental Assay: dHvH pH:4.0 ; Experimental Assay: dCp pH:3.5 ; Experimental Assay: dHcal pH:3.5 ; Experimental Assay: Tm pH:3.5 ; Experimental Assay: dHvH pH:3.5 ; Experimental Assay: dCp pH:3.0 ; Experimental Assay: dHcal pH:3.0 ; Experimental Assay: Tm pH:3.0 ; Experimental Assay: dHvH pH:3.0 ; Experimental Assay: dCp pH:2.5 ; Experimental Assay: dHcal pH:2.5 ; Experimental Assay: Tm pH:2.5 ; Experimental Assay: dHvH pH:2.5 ; Experimental Assay: dCp pH:2.0 ; Experimental Assay: dHcal pH:2.0 ; Experimental Assay: Tm pH:2.0 ; Experimental Assay: dHvH pH:2.0 ; Derived Quantity: ddG_H2O ; Derived Quantity: ddG_H2O ; Derived Quantity: ddG pH:8.0 ; Derived Quantity: ddG pH:7.0 ; Derived Quantity: ddG pH:6.0 ; Derived Quantity: ddG pH:5.0 ; Derived Quantity: ddG pH:4.0 ; Derived Quantity: ddG pH:3.5 ; Derived Quantity: ddG pH:3.0 ; Derived Quantity: ddG pH:2.5 ; Derived Quantity: ddG pH:2.0 ; Derived Quantity: dTm pH:8.0 ; Derived Quantity: dTm pH:7.0 ; Derived Quantity: dTm pH:6.0 ; Derived Quantity: dTm pH:5.0 ; Derived Quantity: dTm pH:4.0 ; Derived Quantity: dTm pH:3.5 ; Derived Quantity: dTm pH:3.0 ; Derived Quantity: dTm pH:2.5 ; Derived Quantity: dTm pH:2.0
Libraries Mutations for sequence EDWLTFQKKHITNTRDVDCDNIMSTNLFHCKDKNTFIYSRPEPVKAICKGIIASKNVLTTSEFYLSDCNVTSRPCKYKLKKSTNKFCVTCENQAPVHFVGVGSC
Sequence Assay Result Units