All three subunits are required for the reconstitution of an active proton channel (F0) of Escherichia coli ATP synthase (F1F0).
All three subunits are required for the reconstitution of an active proton channel (F0) of Escherichia coli ATP synthase (F1F0).
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大肠杆菌 ATP 合酶 (F1F0) 的活性质子通道 (F0) 的重建需要所有三个亚基。
作者：
E. Schneider;K. Altendorf
来自大肠杆菌的ATP合成酶（F1F0）的膜整合，质子易位F0部分由三种亚基a， b和c组成，其化学计量为1:2:10 +/‐1。我们用三氯乙酸（3 M）在pH 8.0下，脱氧胆酸盐（1%）和N -十四烷基- N， N -二甲基- 3 -氨- 1 -丙磺酸盐（Zwittergent 3 - 14, 5%）存在下解离了F0配合物。用洗脱缓冲液中的三氯乙酸（1 M）凝胶过滤分离亚基。用重色谱法和SDS -凝胶电泳检查各组分的均匀性。在整合到磷脂囊泡后，对每个亚基单独以及所有可能的组合进行了H+转运活性和F1结合的测试。一个功能化的H+通道只能通过与原生F0对应的a1b2c10组合来重建。
The membrane‐integrated, proton‐translocating F0 portion of the ATP synthase (F1F0) from Escherichia coli is built up from three kinds of subunits a, b and c with the proposed stoichiometry of 1:2:10 +/‐ 1. We have dissociated the F0 complex by treatment with trichloroacetate (3 M) at pH 8.0, in the presence of deoxycholate (1%) and N‐tetradecyl‐N, N‐dimethyl‐3‐ammonio‐1‐propanesulfonate (Zwittergent 3‐14, 5%). The subunits were separated by gel filtration with trichloroacetate (1 M) included in the elution buffer. The homogeneity of the fractions was checked by rechromatography and SDS‐gel electrophoresis. After integration into phospholipid vesicles each subunit alone as well as all possible combinations were tested for H+ translocating activity and binding of F1. A functional H+ channel could only be reconstituted by the combination a1b2c10 which corresponds to that of native F0.