Binding thermodynamics of metal ions to HIV-1 ribonuclease H domain.

Metal-protein interactions are not necessarily tight in many transient biological processes, such as cellular signaling, enzyme regulation, and molecular recognition. Here, we analyzed the binding thermodynamics and characterized the structural effect of divalent metal ions, i.e. Mn(2+), Zn(2+), and Mg(2+), to the isolated ribonuclease H (RNH) of human immunodeficiency virus (HIV) using isothermal titration calorimetry (ITC) and circular dichroism. The binding thermodynamics of Mg(2+) to RNH was determined using competition ITC experiments, and the binding affinity of Mg(2+) was found to be about 40- and 400-times lower than those of Mn(2+) and of Zn(2+), respectively. The structural analysis showed that Mg(2+) binding had little effect on the thermal stability of RNH, while Zn(2+) and Mn(2+) binding increased the stability. The thermodynamic characteristics of RNH metal binding, compared to intact HIV reverse transcriptase, and a possible mechanism of conformational change induced upon metal ion binding, in correlation with the structure-function relationship, are discussed.