Seelert H, Poetsch A, Rohlfs M, Dencher N A
Institut für Biochemie, Abt. Physikalische Biochemie, Technische Universität Darmstadt, Petersenstr. 22, D-64287 Darmstadt, Germany.
Biochem J. 2000 Feb 15;346 Pt 1(Pt 1):41-4.
n-Dodecyl-beta-D-maltoside was used as a detergent to solubilize the ammonium sulphate precipitate of chloroplast F(O)F(1)-ATP synthase, which was purified further by dye-ligand chromatography. Upon reconstitution of the purified protein complex into phosphatidylcholine/phosphatidic acid liposomes, ATP synthesis, driven by an artificial DeltapH/Deltapsi, was observed. The highest activity was achieved with ATP synthase solubilized in n-dodecyl-beta-D-maltoside followed by chromatography with Red 120 dye. The optimal dye for purification with CHAPS was Green 5. All known subunits were present in the monodisperse proton-translocating ATP synthase preparation obtained from chloroplasts.
使用正十二烷基-β-D-麦芽糖苷作为去污剂来溶解叶绿体F(O)F(1)-ATP合酶的硫酸铵沉淀，该沉淀通过染料配体色谱法进一步纯化。将纯化的蛋白质复合物重构到磷脂酰胆碱/磷脂酸脂质体中后，观察到由人工ΔpH/ΔΨ驱动的ATP合成。用正十二烷基-β-D-麦芽糖苷溶解并随后用Red 120染料进行色谱分离的ATP合酶活性最高。用CHAPS纯化的最佳染料是Green 5。在从叶绿体获得的单分散质子转运ATP合酶制剂中存在所有已知的亚基。
