Molecular determination by electron microscopy of the actin filament end structure
Molecular determination by electron microscopy of the actin filament end structure
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DOI：
10.1016/j.jmb.2006.06.056
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发表时间：
2007-01-12
影响因子：
5.6
通讯作者：
Maeda, Yuichiro
中科院分区：
文献类型：
--
作者：
Narita, Akihiro;Maeda, Yuichiro
在真核细胞中，肌动蛋白丝通过改变其在细胞中的时空分布而发挥各种重要作用。肌动蛋白丝的分布受末端结合蛋白（包括加帽蛋白（肌肉中的CapZ）、Arp 2/3复合物、凝溶胶蛋白、肌动蛋白和原调节蛋白）与肌动蛋白丝末端的结合调节。为了确定这些法规的性质，结构阐明肌动蛋白的毒端结合蛋白复合物是至关重要的。在这里，我们已经开发了新的程序的基础上，单粒子分析，以确定结构的肌动蛋白丝的电子显微镜照片。在这些程序中，肌动蛋白丝图像的极性，以及每个肌动蛋白原聚体的方位角方向和轴向位置在一个短的拉伸附近的丝端，被准确地确定。这大大提高了结构测定的稳定性和准确性。我们测试了我们的程序，从模拟的细丝图像，这是从24个模型结构的肌动蛋白-CapZ复合物的重建结构。这些模型结构是通过将CapZ的原子结构随机对接到肌动蛋白丝的原子模型的倒刺末端而产生的。在24个模型结构中，23个被正确地恢复了本程序。我们发现，我们的分析是强大的对当地的畸变的螺旋扭曲附近的肌动蛋白丝的末端。最后，该程序被成功地应用于确定的肌动蛋白-CapZ复合物的结构，从真实的冷冻电子显微照片的复杂。这是阐明肌动蛋白丝末端详细三维结构的第一种方法。(c)2006爱思唯尔有限公司保留所有权利。
In eukaryotic cells, actin filaments play various crucial roles by altering their spatial and temporal distributions in the cell. The distribution of actin filaments is regulated by the binding of end-binding proteins, including capping protein (CapZ in muscle), the Arp2/3 complex, gelsolin, formin and tropomodulin, to the end of the actin filament. In order to determine the nature of these regulations, structural elucidations of actin filament-end-binding protein complexes are crucially important. Here, we have developed new procedures on the basis of single-particle analysis to determine the structure of the end of actin filaments from electron micrographs. In these procedures, the polarity of the actin filament image, as well as the azimuth orientation and the axial position of each actin protomer within a short stretch near the filament end, were determined accurately. This improved both the stability and accuracy of the structural determination dramatically. We tested our procedures by reconstructing structures from simulated filament images, which were obtained from 24 model structures for the actin-CapZ complex. These model structures were generated by random docking of the atomic structure of CapZ to the barbed end of an atomic model of the actin filament. Of the 24 model structures, 23 were recovered correctly by the present procedures. We found that our analysis was robust against local aberrations of the helical twist near the end of the actin filament. Finally, the procedures were applied successfully to determine the structure of the actin-CapZ complex from real cryo-electron micrographs of the complex. This is the first method for elucidating the detailed 3D structures at the end of the actin filament. (c) 2006 Elsevier Ltd. All rights reserved.