THERMODYNAMICS OF BINDING OF MONONUCLEOTIDES TO RIBONUCLEASE-T1

The binding of mononucleotides to ribonucleases has received much attention in recent years, in part because of the possibility that the different specificities of the ribonucleases might be of interest in connection with specific protein-nucleic acid interactions. Among the most widely studied ribonucleases are ribonuclease A (RNase A), which is specific for pyrimidine nucleotides, RNase U2, specific for purine nucleotides, and RNase T1, specific for guanosine nucleotides (Egami, E.; Oshima, T.; Uchida, T. Mol. Biol. Biochem. Biophys. 1980, 32, 250-277). In this paper we report determinations of the binding constants and enthalpies of binding of several mononucleotides to RNase T1, using both isothermal titration calorimetry (ITC) and differential scanning calorimetry (DSC). ITC was performed at several temperatures to yield values for the heat capacity changes. The DSC results extended the temperature range of validity of many of the thermodynamic data. The binding constants at 25-degrees-C ranged from 5.8 x 10(5) M-1 for 2'-GMP to 7.7 x 10(2) M-1 for 2'-CMP. The enthalpy variation was much smaller, ranging from -9.09 kcal mol-1 for 2'-GMP to -8.22 kcal mol-1 for 2'-CMP.