ISOLATION OF THE MITOCHONDRIAL F1-F0 ADENOSINE-TRIPHOSPHATASE BY SEPHAROSE-HEXYLAMMONIUM CHROMATOGRAPHY - PROPERTIES AND RECONSTITUTION IN LIPOSOMES

Lauryl dimethylamino oxide, a zwitterionic detergent, was employed to solubilize the H+ ATPase from beef heart mitochondria. A simple preparation procedure was devised to obtain F1-F0 based on a method described to purify F1 ATPase which consists of the selective adsorption of F1 to Sepharose-hexylammonium beads. The preparation showed .apprx. 18 bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis; 5 correspond to F1 subunits and the rest probably to the stalk and hydrophobic sector F0. The binding of [14C]dicyclohexylcarbodiimide to a low-molecular-weight component of this preparation was demonstrated. The F1-F0 complex was reconstituted into phospholipid vesicles which displayed ATP-Pi exchange and ATP-dependent 9-aminoacridine fluorescence quenching, both sensitive to proton channel inhibitors.