Selection and Characterization of Artificial Proteins Targeting the Tubulin α Subunit
Selection and Characterization of Artificial Proteins Targeting the Tubulin α Subunit
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DOI：
10.1016/j.str.2018.12.001
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发表时间：
2019-03-05
期刊：
影响因子：
5.7
通讯作者：
Gigant, Benoit
中科院分区：
文献类型：
--
作者：
Campanacci, Valerie;Urvoas, Agathe;Gigant, Benoit
微管是由α - β -微管蛋白异二聚体构成的真核细胞的细胞骨架细丝。非微管小管蛋白的结构研究主要依赖于阻止其自组装的分子，并被用作结晶伴侣。在这里，我们从α Rep文库中鉴定出α微管蛋白特异性的人工蛋白。浊度实验表明，这些α - Reps以剂量依赖的方式阻碍微管组装，全内反射荧光显微镜进一步显示，它们特异性地阻止微管（-）端生长。结构数据表明，它们是通过靶向α -微管蛋白纵向表面来实现的。有趣的是，在所研究的一个复合物中，α亚基的构象介于最常见的微管x射线结构和微管结构之间，强调了微管蛋白的可塑性。这些α -微管蛋白特异性α - rep拓宽了微管动力学及其调控机制研究的工具范围。
Microtubules are cytoskeletal filaments of eukaryotic cells made of alpha beta-tubulin heterodimers. Structural studies of non-microtubular tubulin rely mainly on molecules that prevent its self-assembly and are used as crystallization chaperones. Here we identified artificial proteins from an alpha Rep library that are specific to alpha-tubulin. Turbidity experiments indicate that these alpha Reps impede microtubule assembly in a dose-dependent manner and total internal reflection fluorescence microscopy further shows that they specifically block growth at the microtubule (-) end. Structural data indicate that they do so by targeting the alpha-tubulin longitudinal surface. Interestingly, in one of the complexes studied, the alpha subunit is in a conformation that is intermediate between the ones most commonly observed in X-ray structures of tubulin and those seen in the microtubule, emphasizing the plasticity of tubulin. These alpha-tubulin-specific alpha Reps broaden the range of tools available for the mechanistic study of microtubule dynamics and its regulation.