Determination of the H⁺ /ATP ratio of the H⁺ transport‐coupled reversible chloroplast ATPase reaction by equilibrium studies

Fluorescence quenching of 9‐aminoacridine was followed to ascertain the state of light‐induced transthylakoidal Δ H + at energetic equilibrium with a subsequently added mixture of ATP, ADP and phosphate. In the measured range, the logarithm of relative fluorescence quenching at equilibrium is a linear function of the imposed phosphate potential. The slope of the line is shown by mathematical deduction to be equal to the ATP/H + stoichiometry of the H + ‐coupled reversible ATPase reaction. Determination of the stoichiometry by this method neither relies on the standard phosphate potential nor on the internal thylakoid volume. The results confirm that during ATP synthesis or hydrolysis 3 H + per ATP are translocated through the CF 0 ‐CF 1 complex.