Kleiner Ralph E, Ti Shih-Chieh, Kapoor Tarun M
Laboratory of Chemistry and Cell Biology, The Rockefeller University, New York, New York 10065, USA.
J Am Chem Soc. 2013 Aug 28;135(34):12520-3. doi: 10.1021/ja405199h. Epub 2013 Aug 15.
Microtubules are hollow tube-like biological polymers required for transport in diverse cellular contexts and are important drug targets. Microtubule function depends on interactions with associated proteins and post-translational modifications at specific sites located on its interior and exterior surfaces. However, we lack strategies to selectively perturb or probe these basic biochemical mechanisms. In this work, by combining amber suppression-mediated non-natural amino acid incorporation and tubulin overexpression in budding yeast, we demonstrate, for the first time, a general strategy for site-specific chemistry on microtubules. Probes and labels targeted to precise sites on the interior and exterior surfaces of microtubules will allow analysis and modulation of interactions with proteins and drugs, and elucidation of the functions of post-translational modifications.
微管是一种中空的管状生物聚合物，在多种细胞环境中运输都需要它，并且是重要的药物靶点。微管功能依赖于与相关蛋白质的相互作用以及在其内外表面特定位点的翻译后修饰。然而，我们缺乏选择性干扰或探究这些基本生化机制的策略。在这项工作中，通过结合琥珀抑制介导的非天然氨基酸掺入和芽殖酵母中的微管蛋白过表达，我们首次证明了一种在微管上进行位点特异性化学修饰的通用策略。靶向微管内外表面精确位点的探针和标记物将有助于分析和调节与蛋白质及药物的相互作用，并阐明翻译后修饰的功能。
