Thermal unfolding of monomeric Ca(II),Mg(II)-ATPase from sarcoplasmic reticulum of rabbit skeletal muscle

AbstractThe thermal unfolding of monomeric and delipidated Ca2+-ATPase, solubilized in C12E8, can be appropriately described as a non-two-state irreversible denaturation, with only one endothermic peak. In the Ca2+ concentration range (0–0.5 mM) which stimulates the ATPase activity of solubilized monomeric ATPase, Ca2+ shifts the critical temperature midpoint of the denaturation process (Tm) from 42 to 50°C without segregation of the endothermic peak into two separate components. Because 20 mM Mg2+ only shifts the Tm from 42 to 44°C, we conclude that the effect of Ca2+ upon the Tm is likely to be due to binding to the high affinity Ca2+ sites in the ATPase. The effect of Ca2+ upon the enthalpy of denaturation is biphasic, suggesting the presence of low affinity Ca2+ sites (K0.5 in the millimolar range) in monomeric and solubilized ATPase