Proton Translocation Coupled to ATP Hydrolysis in Rat Liver Mitochondria

Glass electrode measurements of the pH of anaerobic suspensions of rat liver mitochondria during the hydrolysis of ATP to ADP and P i by the oligomycin‐sensitive ATPase show that almost 2H + ions are translocated outwards through the M phase of the cristae membrane per ATP molecule hydrolysed. Adenylate kinase activity is minimised by the presence of EDTA, and valinomycin must be added to the normal mitochondrial suspensions to collapse the membrane potential and permit the outwardly translocated H + ions to diffuse practically quantitatively to the glass electrode system. Classical inhibitor studies of the overall ATPase reaction catalysed by the mitochondria indicate that the specific ATP/ADP porter is not rate‐limiting under the conditions of these experiments; and it is also shown that the overall process of ATP entry and ADP + P i exit through the porter systems in the M phase is not coupled to net proton translocation. It is concluded that the ATPase system responsible for or coupled to proton translocation has a proton translocation quotient (→H + /P) of 2, corresponding to that of the type II ATPase of the chemiosmotic hypothesis.