Structural model of full-length human Ku70-Ku80 heterodimer and its recognition of DNA and DNA-PKcs
Structural model of full-length human Ku70-Ku80 heterodimer and its recognition of DNA and DNA-PKcs
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DOI：
10.1038/sj.embor.7400847
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发表时间：
2007-01-01
期刊：
影响因子：
7.7
通讯作者：
Llorca, Oscar
中科院分区：
文献类型：
--
作者：
Rivera-Calzada, Angel;Spagnolo, Laura;Llorca, Oscar
在非同源末端连接过程中DNA双链断裂的识别是由Ku 70-Ku 80蛋白进行的，Ku 70-Ku 80蛋白是一种150 kDa的异源二聚体，它将DNA修复激酶DNA依赖性蛋白激酶催化亚基（DNA-PKcs）募集到损伤处。确定了截短的Ku 70-Ku 80的原子结构;然而，Ku 80的亚基特异性羧基末端结构域-对于结合DNA-PKcs是必需的-仅在分离中确定，并且Ku 70的C末端结构域在其DNA结合构象中未解析。这两个区域都是保守的，并介导哺乳动物特异性的蛋白质-蛋白质相互作用。在这里，我们重建的三维结构的人全长Ku 70-Ku 80二聚体在25埃分辨率，单独和复杂的DNA，通过使用单粒子电子显微镜。我们绘制了这两个亚基的C-末端区域，以及它们在DNA和DNA-PKcs结合后的构象变化，以在全长Ku 70-Ku 80蛋白的背景下定义这些结构域在DNA修复期间的功能的分子模型。
Recognition of DNA double-strand breaks during nonhomologous end joining is carried out by the Ku70-Ku80 protein, a 150 kDa heterodimer that recruits the DNA repair kinase DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to the lesion. The atomic structure of a truncated Ku70-Ku80 was determined; however, the subunit-specific carboxy-terminal domain of Ku80-essential for binding to DNA-PKcs-was determined only in isolation, and the C-terminal domain of Ku70 was not resolved in its DNA- bound conformation. Both regions are conserved and mediate protein-protein interactions specific to mammals. Here, we reconstruct the three-dimensional structure of the human full-length Ku70-Ku80 dimer at 25 angstrom resolution, alone and in complex with DNA, by using single-particle electron microscopy. We map the C-terminal regions of both subunits, and their conformational changes after DNA and DNA-PKcs binding to define a molecular model of the functions of these domains during DNA repair in the context of full- length Ku70-Ku80 protein.