鉴定能识别组蛋白“尾部”翻译后修饰的蛋白质的方法。
Approach to profile proteins that recognize post-translationally modified histone "tails".
机构信息
Laboratory of Chemistry and Cell Biology, the Rockefeller University, New York, New York 10065, USA.
出版信息
J Am Chem Soc. 2010 Mar 3;132(8):2504-5. doi: 10.1021/ja909741q.
Post-translational modifications (PTMs) of histones, proteins onto which DNA is packaged, are involved in many biological processes, including transcription, recombination, and chromosome segregation. As these PTMs can be dynamic, combinatorial, and mediators of weak interactions, the comprehensive profiling of all proteins that recognize histone PTMs is a daunting task. Here we describe an approach to design probes that can be used to identify proteins that directly interact with modified histones. Protein structure was used to guide the introduction of a photo-cross-linker in the probe, so as to convert weak interactions into covalent linkages. The probe also included an alkyne group to facilitate click chemistry-mediated conjugation of reporter tags for the rapid and sensitive detection (via rhodamine) and affinity enrichment (via biotin) of labeled proteins. In particular, we developed and validated a probe that can selectively capture proteins that recognize trimethyled lysine-4 of histone H3 (H3K4me3) in whole proteomes. A complete profiling of H3K4Me3 binding proteins should shed new light on cellular processes regulated by this PTM.
组蛋白的翻译后修饰（PTMs）是指 DNA 包装的蛋白质上的修饰，参与许多生物学过程，包括转录、重组和染色体分离。由于这些 PTMs 可以是动态的、组合的，并且是弱相互作用的介导物，因此全面分析所有识别组蛋白 PTMs 的蛋白质是一项艰巨的任务。在这里，我们描述了一种设计探针的方法，该探针可用于鉴定与修饰组蛋白直接相互作用的蛋白质。利用蛋白质结构来指导在探针中引入光交联剂，以便将弱相互作用转化为共价键。该探针还包含一个炔基基团，以促进点击化学介导的报告标签的连接，从而快速灵敏地检测（通过罗丹明）和亲和富集（通过生物素）标记的蛋白质。特别是，我们开发并验证了一种探针，该探针可以选择性地捕获识别组蛋白 H3 中赖氨酸-4 三甲基化（H3K4me3）的蛋白质。对 H3K4Me3 结合蛋白的全面分析应该为该 PTM 调控的细胞过程提供新的认识。
Zotero 插件