Evidence for a Structure-Function Relationship in the Activation of Calcineurin by Metal Ions

Kinetic parameters for the metal ion activation of calcineurin at 30°C were found to be correlated to the formation constants (log K1) of the activating metal ions (Mn2+, Co2+, Ni2+, Zn2+, and Mg2+) with the substrate para-nitrophenyl phosphate (pNPP). Metal ion activation measured by kcat/Kact increased with increasing formation constant, although this correlation failed to explain the lack of activation by other metal ions having similar formation constants with pNPP, Ca2+, and Sr2+, for example. An empirical model describing metal ion activation was developed by inclusion of another constant for the metal ions, the hydrolysis constant (pKa M-H2O) of water coordinated to the metal ion with an increase in the pKa related to decreasing activation. The resulting two parameter equation improved the relationship between metal ion properties and enzyme activation. Nonactivating metal ions have the highest values of Ka providing a possible explanation for their failure to cause activation. Derivation of the two-parameter equation inferred that the two properties of metal ions may be important determinants of the role(s) of exogenous metal ion in calcineurin catalysis.