Slow transition of phosphoenzyme from ADP-sensitive to ADP-insensitive forms in solubilized Ca2+, Mg2+-ATPase of sarcoplasmic reticulum: evidence for retarded dissociation of Ca2+ from the phosphoenzyme.

Solubilized Ca²⁺, Mg²⁺-ATPase of sarcoplasmic reticulum was phosphorylated with ATP without added MgCl₂. The phosphoenzyme formed was ADP-sensitive. Ca²⁺ in the medium was chelated after phosphorylation. This induced a slow transition of the phosphoenzyme from ADP-sensitive to ADP-insensitive forms. The ADP-sensitivity was restored by subsequent addition of CaCl₂. These results showed that the transition was caused by dissociation of Ca²⁺ bound to the phosphoenzyme. Further observations indicated that, when Ca²⁺ in the medium was chelated, Ca²⁺ bound to the phosphoenzyme was dissociated much more slowly than Ca²⁺ bound to the dephosphoenzyme. This suggests a possible formation of the occluded form of the Ca²⁺-binding site in the phosphoenzyme.