USE OF HPLC FOR THE STUDY OF ADP BINDING TO CHLOROPLAST ATPase. I. INFLUENCE OF EXPERIMENTAL CONDITIONS AND PROPOSITION OF MECHANISM

Abstract

The binding of ADP to chloroplast ATPase (CF₁) has been studied by chromatographic methods (Hummel and Dreyer method and chromatographic determination of the enzymatic inhibition).

The influence of different factors has been studied: temperature, ionic strength, activation treatments, pyrophosphate fixation. From the comparison of the dependence of ADP binding (and in certain cases, of ATP binding), with that of the enzymatic activity, it is shown that the conclusions drawn for the dissociation constants cannot be extended to the Michaelis constant.

The role of the lysine present in the nucleotide binding sites, already emphasized by several authors, has been proposed to be the ligand, in the protonated form, of the negatively charged phosphate groups of ADP or ATP.

ACKNOWLEDGMENTS

We wish to thank Mrs Jeanine Le Cruer and Mrs Véronique Mary for their skillful assistance in preparing CF₁, and Mrs Monal Paulin for careful typing of this manuscript.