Mitochondrial ATPase Complex

A stable, oligomycin-sensitive, ATPase preparation can be isolated from rat liver mitochondria by refinement of the deoxycholate-solubilization procedure (Soper, J. W., and Pedersen, P. L. (1976) Biochemistry 15, 2682). NADM dehydrogenase, succinic dehydrogenase, and cytochrome oxidase activities cannot be detected by enzymatic assay. Spectral measurements reveal that the content of individual cytochromes is less than 0.05 nmol x mg-’ protein. ATPase activity of the preparation (-13 pmol ATP hydroIyzed/min/mg) is inhibited more than 80% by oligomycin. Electron microscopy of the freshly isolated preparation reveals it to be highly dispersed and membrane-free, and to contain molecules exhibiting a tripartite structural arrangement consisting of the F,-ATPase headpiece, a basepiece, and a stalk connecting these 2 units. The thickness of the basepiece (-60 A) is sufficient to span the hydrophobic phase of the inner mitochondrial membrane. Two types of F,headpieces are present, one of which exhibits the typical spherical structure of membrane-bound F1, and a second of which is more compact and elliptical in appearance. Polyacrylamide gel electrophoresis in sodium dodecyl sulfate reveals the presence of all subunits of FL-ATPase, the oligomycin sensitivity-conferring protein, and several additional polypeptides which are candidates for the basepiece. When bound to soy bean phospholipid vesicles the preparation catalyzes an oligomycin-sensitive ATP-Pi exchange reaction in the absence of other protein-coupling factors. This report summarizes the first description of the properties of an oligomycin-sensitive ATPase preparation from rat liver, and the first description of an energy-transducing ATPase preparation in which individual molecules in dispersed form are seen to exhibit clearly a tripartite headpiece-stalk-basepiece arrangement.