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

Abstract

The H+‐ATPase from chloroplasts (CF0F1) was isolated, purified and reconstituted into liposomes from phosphatidylcholine/phosphatidic acid. A transmembrane pH difference, pH, and a transmembrane electric potential difference, Ψ, were generate by an acid/base transition. The rate of ATP synthesis was measured at constant pH and constant Ψ 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 pH = 1.8. The dependence of the rate of ATP synthesis on ADP concentration was measured with CF0F1 in the oxidized state, Eox, and in the reduced state, Ered. The results can be described by Michaelis‐Menten kinetics with the following parameters: Vmax= 0.5 s−1, Km= 8 μLM for Eox and Vmax= 2.0 s−1, Km= 8 μLM for Ered. Copyright © 1992, Wiley Blackwell. All rights reserved