Spontaneous reactivation of covalently labeled proton adenosinetriphosphatase.

Bovine heart mitochondrial adenosinetriphosphatase selectively labeled by [14C]-N,N'-dicyclohexylcarbodiimide or [14C]-7-chloro-4-nitro-2,1,3-benzoxadiazole was used together with other components to form reconstituted submitochondrial particles. When assayed for ATP hydrolysis under normal hydrolysis condition, these labeled submitochondrial particles were found to increase slowly in specific activity with preincubation time, without losing the covalent label. But when assayed for oxidative phosphorylation, the ratio of the specific activity of the same labeled particles to that of the control particles was higher and was unaffected by preincubation. If the labeled particles had been treated by a simulated procedure for oxidative phosphorylation measurement before the ATPase assay, their specific activities for ATP hydrolysis were also found to be higher and unaffected by preincubation. These observations are difficult to reconcile with the alternating three-site model for proton adenosinetriphosphatase or any model which requires the sequential action of three identical sites for ATP hydrolysis and synthesis. A new model with one active and two latent interacting sites is proposed for interpreting the present data.