Chemical proteomics enables the global analysis of small molecule-protein interactions in native biological systems and has emerged as a versatile approach for ligand discovery. The range of small molecules explored by chemical proteomics has, however, remained limited. Here, we describe a diversity-oriented synthesis (DOS)-inspired library of stereochemically defined compounds bearing diazirine and alkyne units for UV light-induced covalent modification and click chemistry enrichment of interacting proteins, respectively. We find that these “photo-stereoprobes” interact in a stereoselective manner with hundreds of proteins from various structural and functional classes in human cells and demonstrate that these interactions can form the basis for high-throughput screening-compatible NanoBRET assays. Integrated phenotypic screening and chemical proteomics identified photo-stereoprobes that modulate autophagy by engaging the mitochondrial serine protease CLPP. Our findings show the utility of DOS-inspired photo-stereoprobes for expanding the ligandable proteome, furnishing target engagement assays, and facilitating the discovery and characterization of bioactive compounds in phenotypic screens.

中文翻译：

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扩展配体蛋白质组的化学工具：面向多样性的基于合成的光反应立体探针


化学蛋白质组学能够对天然生物系统中的小分子-蛋白质相互作用进行整体分析，并已成为一种多功能的配体发现方法。然而，化学蛋白质组学探索的小分子范围仍然有限。在这里，我们描述了一个面向多样性的合成 （DOS） 启发的立体化学定义的化合物库，这些化合物带有二氮嗪和炔烃单元，分别用于紫外光诱导的共价修饰和相互作用蛋白质的点击化学富集。我们发现这些“光立体探针”以立体选择性方式与人类细胞中来自不同结构和功能类别的数百种蛋白质相互作用，并证明这些相互作用可以构成高通量筛选兼容的 NanoBRET 检测的基础。综合表型筛选和化学蛋白质组学鉴定了通过参与线粒体丝氨酸蛋白酶 CLPP 来调节自噬的光立体探针。我们的研究结果表明，DOS 启发的光立体探针可用于扩展可配体蛋白质组、提供靶标参与测定以及促进表型筛选中生物活性化合物的发现和表征。