The identification and validation of a small molecule’s targets is a major bottleneck in the discovery process for tuberculosis antibiotics. Activity-based protein profiling (ABPP) is an efficient tool for determining a small molecule’s targets within complex proteomes. However, how target inhibition relates to biological activity is often left unexplored. Here, we study the effects of 1,2,3-triazole ureas on Mycobacterium tuberculosis (Mtb). After screening ∼200 compounds, we focus on 4 compounds that form a structure-activity series. The compound with negligible activity reveals targets, the inhibition of which is functionally less relevant for Mtb growth and viability, an aspect not addressed in other ABPP studies. Biochemistry, computational docking, and morphological analysis confirms that active compounds preferentially inhibit serine hydrolases with cell wall and lipid metabolism functions and that disruption of the cell wall underlies biological activity. Our findings show that ABPP identifies the targets most likely relevant to a compound's antibacterial activity.

小分子靶点的识别和验证是结核病抗生素发现过程中的主要瓶颈。基于活性的蛋白质分析 (ABPP) 是确定复杂蛋白质组中小分子目标的有效工具。然而，靶点抑制与生物活性的关系通常未被探索。在这里，我们研究了 1,2,3-三唑脲对结核分枝杆菌( Mtb ) 的影响。在筛选了 200 种化合物后，我们重点研究了 4 种形成构效系列的化合物。活性可忽略不计的化合物揭示了靶点，其抑制在功能上与Mtb的相关性较低生长和生存能力，这是其他 ABPP 研究中未涉及的一个方面。生物化学、计算对接和形态分析证实，活性化合物优先抑制具有细胞壁和脂质代谢功能的丝氨酸水解酶，并且细胞壁的破坏是生物活性的基础。我们的研究结果表明，ABPP 确定了最可能与化合物的抗菌活性相关的目标。