探索硫酸二甲酯用于体内蛋白质组足迹
Exploring dimethylsulfate for in vivo proteome footprinting
摘要：
蛋白质足迹法是一种新的方法，其基于探测，通常使用质谱法，不同氨基酸残基对修饰试剂的反应性。由此获得的数据使人们能够对蛋白质构象及其分子间相互作用进行推断。迄今为止，大多数蛋白质足迹研究都是在体外对单个蛋白质进行的。我们探索是否有可能采用类似的方法与活细胞内的蛋白质，采用硫酸二甲酯（DMS），一种广泛用于核酸体内足迹的试剂。DMS可诱导蛋白质上赖氨酸、组氨酸和谷氨酸残基的甲基化。使用模型的组蛋白H2 B/H2 AZ异二聚体组装在体外和从染色质在体内处理，我们表明，由氘代DMS的甲基化允许一个区分特定的残基的可访问性和蛋白质的环境/结构背景。检测蛋白质构象的变化或它们在体内的相互作用可以提供一种新的方法来鉴定参与各种细胞内途径的蛋白质，并有助于寻找有前景的药物靶点和疾病的生物标志物。
英文摘要：
Protein footprinting is a new methodology that is based on probing, typically with the use of mass spectrometry, of reactivity of different aminoacid residues to a modifying reagent. Data thus obtained allow one to make inferences about protein conformations and their intermolecular interactions. Most of the protein footprinting studies so far have been performed on individual proteins in vitro. We explore whether a similar approach is possible with the proteins inside of living cells, employing dimethylsulfate (DMS), a reagent widely used for the in vivo footprinting of nucleic acids. DMS can induce methylation of the lysine, histidine and glutamate residues on proteins. Using models of the histone H2B/H2AZ heterodimer assembled in vitro and from chromatin treated in vivo, we show that the methylation by deuterated DMS allows one to distinguish the accessibility of a particular residue in and out of the protein's environmental/structural context. The detection of changes in protein conformations or their interactions in vivo can provide a new approach to the identification of proteins involved in various intracellular pathways and help in the search for perspective drug targets and biomarkers of diseases.
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