Adenosine triphosphatase from rat liver mitochondria: Separate sites involved in ATP hydrolysis and in the reversible, high affinity binding of ADP

Abstract

Homogeneous ATPase from rat liver mitochondria binds one mole of ADP per mole of enzyme reversibly, and with high affinity (K D = 1–2 μM). The high affinity binding site is highly specific for ADP and dADP. AMP does not bind. Agents which inhibit ATP hydrolysis have little inhibitory effect on the high affinity binding of ADP. These agents include adenylyl imidodiphosphate (AMP-PNP), azide, sucrose, and the divalent cation Mg ++. AMP-PNP inhibits ATPase activity in phosphorylating membrane preparations of rat liver mitochondria by about 90 percent, but is without effect on ATP synthesis. These results are consistent with the view that the purified soluble, and the membrane-bound ATPase of rat liver mitochondria contain separate sites involved in ATP hydrolysis and in the reversible, high affinity binding of ADP.