Localized reconstruction of subunits from electron cryomicroscopy images of macromolecular complexes.
Localized reconstruction of subunits from electron cryomicroscopy images of macromolecular complexes.
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DOI：
10.1038/ncomms9843
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发表时间：
2015-11-04
影响因子：
16.6
通讯作者：
Huiskonen JT
中科院分区：
文献类型：
--
作者：
Ilca SL;Kotecha A;Sun X;Poranen MM;Stuart DI;Huiskonen JT
电子低温显微镜可以产生高度有序的大分子复合物的近原子分辨率结构。然而，通常一些亚基以灵活的方式结合，具有与复合物的其余部分不同的对称性，或者以亚化学计量的量存在，限制了可达到的分辨率。在这里，我们报告了一个一般的方法，这样的亚基的局部三维重建。在确定颗粒取向之后，可以提取对应于子单元的局部区域并将其作为单个颗粒处理。我们演示了使用三个例子，包括灵活的组装和复杂的窝藏亚基与部分占用或不匹配的对称性的方法。最值得注意的是，该方法允许在病毒衣壳内的三重对称轴处结合的单体RNA依赖性RNA聚合酶的准确拟合，首次揭示了其确切的方向和与衣壳蛋白的相互作用。局部重建有望在一系列具有挑战性的生物系统中提供新的生物学见解。 电子低温显微镜可以允许大分子结构的阐明;然而，不同组分之间的对称性不匹配限制了可达到的分辨率。在这里，作者提出了一种从这些组件中提取和保留信息的计算方法。
Electron cryomicroscopy can yield near-atomic resolution structures of highly ordered macromolecular complexes. Often however some subunits bind in a flexible manner, have different symmetry from the rest of the complex, or are present in sub-stoichiometric amounts, limiting the attainable resolution. Here we report a general method for the localized three-dimensional reconstruction of such subunits. After determining the particle orientations, local areas corresponding to the subunits can be extracted and treated as single particles. We demonstrate the method using three examples including a flexible assembly and complexes harbouring subunits with either partial occupancy or mismatched symmetry. Most notably, the method allows accurate fitting of the monomeric RNA-dependent RNA polymerase bound at the threefold axis of symmetry inside a viral capsid, revealing for the first time its exact orientation and interactions with the capsid proteins. Localized reconstruction is expected to provide novel biological insights in a range of challenging biological systems. Electron cryomicroscopy can allow the elucidation of macromolecular structures; however, mismatches in symmetry between different components limit the attainable resolution. Here, the authors set out a computational method for extracting and retaining information from such components.