Mitochondrial ATP synthasome -: Three-dimensional structure by electron microscopy of the ATP synthase in complex formation with carriers for Pi and ADP/ATP
Mitochondrial ATP synthasome -: Three-dimensional structure by electron microscopy of the ATP synthase in complex formation with carriers for Pi and ADP/ATP
复制标题
DOI：
10.1074/jbc.m401353200
复制
发表时间：
2004-07-23
影响因子：
4.8
通讯作者：
Pedersen, PL
中科院分区：
文献类型：
--
作者：
Chen, C;Ko, Y;Pedersen, PL
在线粒体中制造ATP的最终步骤需要ATP合成酶（F0F1），该合成酶由两个马达组成，一个磷酸载体（PIC）和一个腺嘌呤核苷酸载体（ANC）。在温和的条件下，这些实体会分解成ATP合成酶/PIC/ANC复合物或“ATP合成体”(Ko， Y. H., Delannoy， M, Hullihen， J, Chiu， W., and Pedersen， P. L.（2003）。化学，278,12305 - 12309)。为了获得这一大型复合物或“代谢物”的三维信息以及PIC和ANC在其中的位置，我们将ATP合体分散到单个复合物中，并通过电子显微镜（EM）观察负染色图像，清楚地显示了经典的头部、中心柄和基段。平行免疫电镜研究显示PIC和ANC位于基片段的非中心位置，其他研究暗示ATP合成酶/PIC/ANC的化学计量接近1:1。单个ATP合体图像（7506）被装箱，并使用EMAN软件，以23埃的分辨率获得三维模型。值得注意的是，基片段是长方形的，包含两个结构域，其中较大的连接到中心茎，而较小的则作为延伸。与已知结构的对接研究以及免疫- em研究表明，PIC或ANC可能位于较小的区域，而其他转运蛋白则位于较大的区域附近。总的来说，这些发现支持了一种机制，即底物ADP和Pi进入线粒体，F-1上ATP的合成以及ATP的释放和退出是非常局部和高度协调的事件。
The terminal steps involved in making ATP in mitochondria require an ATP synthase (F0F1) comprised of two motors, a phosphate carrier (PIC), and an adenine nucleotide carrier (ANC). Under mild conditions, these entities sub-fractionate as an ATP synthase/PIC/ANC complex or "ATP synthasome" (Ko, Y. H., Delannoy, M, Hullihen, J., Chiu, W., and Pedersen, P. L. ( 2003) J. Biol. Chem. 278, 12305 - 12309). As a first step toward obtaining three-dimensional information about this large complex or "metabolon" and the locations of PIC and ANC therein, we dispersed ATP synthasomes into single complexes and visualized negatively stained images by electron microscopy ( EM) that showed clearly the classical headpiece, central stalk, and basepiece. Parallel immuno-EM studies revealed the presence of PIC and ANC located non-centrally in the basepiece, and other studies implicated an ATP synthase/PIC/ANC stoichiometry near 1: 1: 1. Single ATP synthasome images ( 7506) were boxed, and, using EMAN software, a three-dimensional model was obtained at a resolution of 23 Angstrom. Significantly, the basepiece is oblong and contains two domains, the larger of which connects to the central stalk, whereas the smaller appears as an extension. Docking studies with known structures together with the immuno-EM studies suggest that PIC or ANC may be located in the smaller domain, whereas the other transporter resides nearby in the larger domain. Collectively, these finding support a mechanism in which the entry of the substrates ADP and Pi into mitochondria, the synthesis of ATP on F-1, and the release and exit of ATP are very localized and highly coordinated events.