Kinetics of ATP synthesis catalyzed by the H⁺‐ATPase from chloroplasts (CF₀F₁) reconstituted into liposomes and coreconstituted with bacteriorhodopsin

The H⁺-ATPase from chloroplasts (CF₀F₁) was isolated, purified and reconstituted into liposomes from phosphatidylcholine/phosphatidic acid. A transmembrane pH difference, ΔpH, and a transmembrane electric potential difference, ΔΨ, were generate by an acid/base transition. The rate of ATP synthesis was measured at constant ΔpH and constant ΔΨ as a function of temperature between 5°C and 45°C. The activation energy was 55 kJ mol⁻¹.

CF₀F₁ was coreconstituted with bacteriohodopsin at a molar ratio of approximately 1:170 in the same type of liposomes. Illumination of the proteoliposomes leads to proton transport into the vesicles generating a constant ΔpH = 1.8. The dependence of the rate of ATP synthesis on ADP concentration was measured with CF₀F₁ in the oxidized state, E^ox, and in the reduced state, E^red. The results can be described by Michaelis-Menten kinetics with the following parameters: V_max= 0.5 s⁻¹, K_m= 8 µLM for E^ox and V_max= 2.0 s⁻¹, K_m= 8 µLM for E^red.