THERMODYNAMIC ANALYSIS OF CALCIUM AND MAGNESIUM BINDING TO CALMODULIN

TO elucidate some aspects still debated concerning the interaction of Ca2+ and Mg2+ with CaM, the thermodynamic binding parameters of Ca2+-C aM and Mg2+-CaM complexes were characterized by flow dialysis and isot hermal microcalorimetry under different experimental conditions. In pa rticular, the enthalpy and entropy changes associated with Ca2+ and Mg 2+ binding to their sites were determined, allowing a better understan ding of the mechanism underlying cation-CaM interactions. Ca2+-CaM int eraction follows an enthalpy-entropy compensation relationship, sugges ting that CaM explores a subspace of isoenergetical conformations whic h is modified by Ca2+ binding. This Ca2+-induced change in CaM dynamic s is proposed to play a key role in CaM function, i.e. in its interact ion with and/or activation of target proteins. Furthermore, data show that Mg2+ does not act as a direct competitor for Ca2+ binding on the four main Ca2+ binding sites, but rather as an allosteric effector. Th is implies that the four main Mg2+ binding sites are distinct from the EF-hand Ca2+ binding sites. Finally, Ca2+ is shown to interact with a uxiliary binding sites on CaM. These weak affinity sites were thermody namically characterized. The results presented here challenge the curr ent accepted view of CaM ion binding.