Cooperativity between the enzymatic sites of F1-ATPase revisited by the use of HPLC methods

The fundamental question of the cooperativity between the enzymatic sites of F1-ATPase is examined in the light of new measurements of the enzymatic rate of ATP hydrolysis by CF1, the enzyme isolated from spinach chloroplasts. The experimental data, obtained with a chromatographic method, fit a model that involves two kinds of independent enzymatic sites working with metal-free ATP, with no need of cooperativity between the sites. Binding measurements between ADP or ATP and CF1 by the chromatographic method of Hummel and Dreyer (1962) also support this conclusion. The present data and interpretation are in agreement with those reported recently (Reynafarje and Pedersen, 1996) which show that the first order rate constant of ATP hydrolysis by MF1, the analogous enzyme from mitochondria, is virtually constant under experimental conditions involving either unisite or multisite hydrolysis of ATP. The present data and interpretation are discussed together with those reported previously, in particular with regard to the methods that were used to support the commonly accepted opposite viewpoint.