The highly purified soluble ATP synthetase complex from mitochondria, containing energy-transfer Factor A (the terminal ADP phosphorylation enzyme of oxidative phosphorylation) and Factor D, catalyzes ATP-P(i) and ATP-ADP exchange reactions. The ATP-P(i) exchange activity is inhibited by low concentrations of the uncouplers of oxidative phosphorylation, oligomycin and p-chloromercnriphenylsulfonate. It is stimulated threefold by dithiothreitol and is Mg(++) dependent. Antiserum to coupling factor 1 (F(1)) also inhibits the ATP-P(i) exchange. The ATP-ADP exchange activity appears to be greater than the ATP-P(i) exchange activity. The results suggest that the nonphosphorylated high-energy intermediate (X approximately C), and possibly the phosphorylated intermediate (X approximately P), are formed on the synthetase. Sites of uncoupler and oligomycin action reside in the terminal ATP synthetase.