Proton ATPase of rat liver mitochondria: A rapid procedure for purification of a stable, reconstitutively active F₁ preparation using a modified chloroform method

A method is described for the purification of rat liver F₁-ATPase by a modification of the chloroform extraction procedure originally described by Beechey et al. (Biochem. J. (1975) 148, 533). Purified liver membrane vesicles are extracted with chloroform in the presence of ATP and EDTA. The procedure yields pure F₁ in only 2-3 h without the necessity of ion-exchange chromatography. The enzyme exhibits the α, β, γ, δ, and ε{lunate} bands characteristic of F₁-ATPase. It has a high ATPase specific activity, and is reconstitutively active, catalyzing high rates of ATP synthesis. Significantly, it can be readily crystallized. If desired, the enzyme can be passed over a gel filtration column to place it in a stabilizing phosphate-EDTA buffer, lyophilized and stored indefinitely at -20°C.