Modulation of the Ca 2+,Mg2+-ATPase of sarcoplasmic reticulum by the hypothalamic hypophyseal inhibitory factor

AbstractWe have studied the effect of the endogenous inhibitor of the Na+ and Ca 2+ pumps, HHIF, on sarcoplasmic reticulum (SR) vesicles. The effect of HHIF on the SR Ca 2+,Mg2+-ATPase activity shows a biphasic pattern. Low HHIF concentrations activate the Ca 2+,Mg2+-ATPase by dissipation of Ca 2+ gradient across the SR membrane. Higher concentrations irreversibly inhibit this activity following a slow kinetic process both in intact SR membranes and in purified Ca 2+,Mg2+-ATPase. Differential scanning calorimetry shows that the Ca 2+,Mg2+-ATPase is denatured after incubation with HHIF concentrations which produced full inhibition of its activity. Micromolar Ca 2+ and millimolar Mg t+ADP protect against the irreversible inhibition of the Ca 2+,Mg2+-ATPase by HHIF. The concentration of HHIF which produces 50% inhibition depends upon SR membrane concentration and upon the lipid:protein ratio in purified Ca 2+,Mg2+-ATPase. From this we have obtained a partition coefficient for binding of HHIF to SR membranes of 0.6 (μg SR protein/ml)−1