Selective Reconstitution of the Tonoplast H + ‐ATPase of the Crassulacean‐Acid Metabolism Plant Kalanchoë daigremontiana

Summary IA detergent removal technique was used to reconstitute solubilized tonoplast proteins of mesophyll cells of the CAM plant Kalanchoe daigremontiana into phosphatidylcholine liposomes. The proteoliposomes were able to hydrolyse ATP and to pump protons across the vesicle membrane. Both activities were inhibited by nitrate, an inhibitor of V-type ATPases. Freeze-fracture micrographs confirmed the incorporation of membrane proteins into liposomes. Increase of specific ATP-hydrolysis activity compared to solubilized tonoplast proteins and SDS-PAGE analysis of reconstituted proteins in comparison with the polypeptide pattern of the purified tonoplast H+-ATPase from the same plant source indicated a highly selective reconstitution of the tonoplast H+-ATPase.