As an anti-tuberculosis target, DprE1 contains two flexible loops (Loop I and Loop II) which have never been exploited for developing DprE1 inhibitors. Here Leu317 in Loop II was discovered as a new functional site to combat drug-resistance in Mycobacterium strains. Based on TCA1, LZDT1 was designed to optimize the hydrophobic interaction with Leu317. A subsequent biochemical and cellular assay displayed increased potency of LZDT1 in inhibiting DprE1 and killing drug-sensitive/-resistant Mycobacterium strains. The improved activity of LZDT1 and its analogue LZDT2 against multidrug resistant tuberculosis was particularly highlighted. For LZDT1, its enhanced interaction with Leu317 also impaired the drug-insensitivity of DprE1 caused by Cys387 mutation. A new nonbenzothiazole lead (LZDT10) with reduced Cys387-dependence was further produced by optimizing interactions with Leu317, improvement directions for LZDT10 were discussed as well. Our research underscores the value of potential functional sites in disordered loops, and affords a feasible way to develop these functional sites into opportunities for drug-resistance management.

作为一种抗结核病靶点，DprE1 包含两个灵活的环（Loop I 和 Loop II），它们从未被用于开发 DprE1 抑制剂。在这里，Loop II 中的 Leu317 被发现是一个新的功能位点，可以对抗分枝杆菌菌株的耐药性。基于 TCA1，LZDT1 旨在优化与 Leu317 的疏水相互作用。随后的生化和细胞测定显示 LZDT1 在抑制 DprE1 和杀死药物敏感/耐药分枝杆菌方面的效力增加菌株。特别强调了 LZDT1 及其类似物 LZDT2 对耐多药结核病的改善活性。对于 LZDT1，其增强的与 Leu317 的相互作用也削弱了由 Cys387 突变引起的 DprE1 的药物不敏感性。通过优化与 Leu317 的相互作用，进一步产生了降低 Cys387 依赖性的新型非苯并噻唑先导 (LZDT10)，并讨论了 LZDT10 的改进方向。我们的研究强调了无序循环中潜在功能位点的价值，并提供了一种可行的方法将这些功能位点开发成耐药性管理的机会。