Study of the mitochondrial phosphate carrier in the course of calcium phosphate accumulation: A requirement for Mg 2+ and ADP of its sensitivity to thiol reagents

Abstract

1. The accumulation of calcium phosphate driven by succinate oxidation is ADP-dependent. In its absence the accumulation stops after a short incubation time and the oxygen uptake is permanently stimulated. This uncoupled oxygen uptake is insensitive to the inhibitors of phosphate transport, like mersalyl and N-ethylmaleimide. When ADP plus Mg 2+ are added to the medium, or when ADP is added in the initial presence of magnesium, the inhibitory action of the thiol reagents on oxygen uptake is re-established. ADP alone or Mg 2+ alone are without any effect. 2. Phosphate/phosphate exchange has been studied, in the absence of ADP, when calcium phosphate accumulation had stopped and oxygen uptake is uncoupled. Under these conditions the exchange process becomes insensitive to thiol reagents. Sensitivity is recovered solely in the presence of ADP plus Mg 2+. 3. When mitochondrial swelling is studied according to the method of Chappell, it also appears that the phosphate carrier loses it sensitivity to mersalyl in the absence of ADP, which confirms the data obtained with phosphate/phosphate exchange experiments. When ADP plus Mg 2+ are added (or present), together with mersalyl, the action of the thiol inhibitor is recovered. ADP and magnesium are inactive separately. EGTA plus Mg 2+ (but not EGTA plus ADP) may substitute for ADP plus Mg 2+ in this process. 4. A possible interaction between the magnesium binding site and the phosphate carrier is considered and discussed.