| Abstract: | Magnesium stimulates phosphorylation of the calcium pump protein of the sarcoplasmic reticulum by inorganic phosphate, but the effect is reversed by high Mg2+]. This reversal is readily explained in terms of the generally accepted existence of two conformational states of the enzyme, E1 and E2. E2 is the form of the enzyme that can be phosphorylated by Pi, and it has one binding site for Mg2+. E1 is the form of the enzyme that has two high-affinity Ca2+ binding sites, and it is phosphorylated by ATP when Ca2+ is bound. Mg2+ can bind weakly to the two Ca2+ sites and to a third site known to be present on E1; this stabilizes E1 at the expense of E2 when Mg2+] is large. Stabilization of E1 at pH 6.2 and 25 degrees C was found to be a highly cooperative function of Mg2+] and was not prevented by increasing Pi]. The latter result requires the existence of a binding site for Pi on E1, with an affinity for Pi comparable to that of E2. Cooperativity with respect to Mg2+] requires that E2 is the stable state of the enzyme in the absence of ligands, with an equilibrium constant E2]/E1] on the order of 10(3) or higher at pH 6.2 and 25 degrees C. |
