Department of Chemistry, University of California at Riverside, Riverside, California.
Department of Biochemistry, University of California at Riverside, Riverside, California.
Protein Sci. 2022 Feb;31(2):432-442. doi: 10.1002/pro.4236. Epub 2021 Nov 26.
Antibiotic resistance is a continually growing challenge in the treatment of various bacterial infections worldwide. New drugs and new drug targets are necessary to curb the threat of infectious diseases caused by multidrug-resistant pathogens. The tryptophan biosynthesis pathway is essential for bacterial growth but is absent in higher animals and humans. Drugs that can inhibit the bacterial biosynthesis of tryptophan offer a new class of antibiotics. In this work, we combined a structure-based strategy using in silico docking screening and molecular dynamics (MD) simulations to identify compounds targeting the α subunit of tryptophan synthase with experimental methods involving the whole-cell minimum inhibitory concentration (MIC) test, solution state NMR, and crystallography to confirm the inhibition of L-tryptophan biosynthesis. Screening 1,800 compounds from the National Cancer Institute Diversity Set I against α subunit revealed 28 compounds for experimental validation; four of the 28 hit compounds showed promising activity in MIC testing. We performed solution state NMR experiments to demonstrate that a one successful inhibitor, 3-amino-3-imino-2-phenyldiazenylpropanamide (Compound 1) binds to the α subunit. We also report a crystal structure of Salmonella enterica serotype Typhimurium tryptophan synthase in complex with Compound 1 which revealed a binding site at the αβ interface of the dimeric enzyme. MD simulations were carried out to examine two binding sites for the compound. Our results show that this small molecule inhibitor could be a promising lead for future drug development.
抗生素耐药性是全球各种细菌感染治疗中不断增长的挑战。为了遏制多药耐药病原体引起的传染病威胁，需要新的药物和新的药物靶点。色氨酸生物合成途径对细菌生长至关重要，但在高等动物和人类中不存在。能够抑制细菌色氨酸生物合成的药物提供了一类新的抗生素。在这项工作中，我们结合了基于结构的策略，使用计算机对接筛选和分子动力学 (MD) 模拟来识别靶向色氨酸合酶α亚基的化合物，并使用涉及全细胞最小抑菌浓度 (MIC) 测试、溶液状态 NMR 和晶体学的实验方法来确认 L-色氨酸生物合成的抑制。对来自国家癌症研究所多样性集 I 的 1800 种化合物进行筛选，针对α亚基发现了 28 种化合物用于实验验证；28 种命中化合物中有 4 种在 MIC 测试中表现出有希望的活性。我们进行了溶液状态 NMR 实验，证明了一种成功的抑制剂，3-氨基-3-亚氨基-2-苯基亚氨基丙酰胺（化合物 1）与α亚基结合。我们还报告了一个与化合物 1 结合的沙门氏菌 enterica 血清型 Typhimurium 色氨酸合酶的晶体结构，该结构揭示了二聚酶的αβ界面上的一个结合位点。进行了 MD 模拟以检查该化合物的两个结合位点。我们的结果表明，这种小分子抑制剂可能是未来药物开发的有前途的先导化合物。
