Simple and rapid purification of F1-ATPase from bovine heart mitochondria by affinity chromatography

A simple and rapid method for the isolation of bovine heart mitochondrial adenosine 5'-triphosphatase (F1-ATPase) was developed. Mitochondria were purified by differential centrifugation and stored frozen. After thawing. F1-ATPase was released by treatment with chloroform. Purification of the enzyme was achieved by polyethylene glycol precipitation followed by chromatography on Procion Navy H-ER beaded cellulose in the presence of MgCl2. F1-ATPase was eluted by ATP in the absence of MgCl2. The purity of the enzyme was proved by SDS-polyacrylamide-gel electrophoresis. The purified F1-ATPase showed slightly non-hyperbolic kinetics towards ATP and nearly complete inhibition in the presence of millimolar concentrations of ADP.