Identifying the target proteins of bioactive small molecules is a key step in understanding mode-of-action of the drug and addressing the underlying mechanisms responsible for a particular phenotype. Proteomics has been successfully used to elucidate the target protein profiles of unmodified and ligand-modified bioactive small molecules. In the latter approach, compounds can be modified via click chemistry and combined with activity-based protein profiling. Target proteins are then enriched by performing a pull-down with the modified ligand. Methods that utilize unmodified bioactive small molecules include the cellular thermal shift assay, thermal proteome profiling, stability of proteins from rates of oxidation, and the drug affinity responsive target stability (DARTS) determination (or read-out). This review highlights recent proteomic approaches utilizing data-dependent analysis and data-independent analysis to identify target proteins by DARTS. When combined with liquid chromatography/tandem mass spectrometry, DARTS enables the identification of proteins that bind to drug molecules that leads to a conformational change in the target protein(s). In addition, an effective strategy is proposed for selecting the target protein(s) from within the pool of analyzed candidates. With additional complementary methods, the biologically relevant target proteins that bind to the small bio-active molecules can be further validated.

中文翻译：

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用药物亲和力响应靶标稳定性和液相色谱/串联质谱分析未修饰生物活性分子的蛋白靶标。

识别具有生物活性的小分子的靶蛋白是理解药物作用方式和解决引起特定表型的潜在机制的关键步骤。蛋白质组学已成功用于阐明未修饰和配体修饰的生物活性小分子的靶蛋白谱。在后一种方法中，化合物可以通过点击化学进行修饰，并与基于活性的蛋白质谱相结合。然后通过用修饰的配体进行下拉来富集靶蛋白。利用未经修饰的生物活性小分子的方法包括细胞热位移分析，热蛋白质组分析，氧化速率产生的蛋白质稳定性以及药物亲和力响应靶标稳定性（DARTS）测定（或读出）。这篇综述重点介绍了利用蛋白质依赖分析和数据独立分析通过DARTS鉴定目标蛋白的最新蛋白质组学方法。当与液相色谱/串联质谱联用时，DARTS可以鉴定与药物分子结合的蛋白质，从而导致目标蛋白质的构象变化。此外，提出了一种有效的策略，用于从分析的候选对象库中选择目标蛋白。使用其他补充方法，可以进一步验证与小生物活性分子结合的生物学相关靶蛋白。DARTS能够鉴定与药物分子结合的蛋白质，从而导致目标蛋白质的构象发生变化。此外，提出了一种有效的策略，用于从分析的候选对象库中选择目标蛋白。使用其他补充方法，可以进一步验证与小生物活性分子结合的生物学相关靶蛋白。DARTS能够鉴定与药物分子结合的蛋白质，从而导致目标蛋白质的构象发生变化。此外，提出了一种有效的策略，用于从分析的候选对象库中选择目标蛋白。使用其他补充方法，可以进一步验证与小生物活性分子结合的生物学相关靶蛋白。