Molodtsov Vadim, Scharf Nathan T, Stefan Maxwell A, Garcia George A, Murakami Katsuhiko S
Department of Biochemistry and Molecular Biology, The Center for RNA Molecular Biology, The Pennsylvania State University, University Park, PA, 16802, USA.
Department of Medicinal Chemistry, College of Pharmacy, University of Michigan, 428 Church St, Ann Arbor, MI, 48109-1065, USA.
Mol Microbiol. 2017 Mar;103(6):1034-1045. doi: 10.1111/mmi.13606. Epub 2017 Jan 10.
Since 1967, Rifampin (RMP, a Rifamycin) has been used as a first line antibiotic treatment for tuberculosis (TB), and it remains the cornerstone of current short-term TB treatment. Increased occurrence of Rifamycin-resistant (RIF ) TB, ∼41% of which results from the RpoB S531L mutation in RNA polymerase (RNAP), has become a growing problem worldwide. In this study, we determined the X-ray crystal structures of the Escherichia coli RNAPs containing the most clinically important S531L mutation and two other frequently observed RIF mutants, RpoB D516V and RpoB H526Y. The structures reveal that the S531L mutation imparts subtle if any structural or functional impact on RNAP in the absence of RIF. However, upon RMP binding, the S531L mutant exhibits a disordering of the RIF binding interface, which effectively reduces the RMP affinity. In contrast, the H526Y mutation reshapes the RIF binding pocket, generating significant steric conflicts that essentially prevent any RIF binding. While the D516V mutant does not exhibit any such gross structural changes, certainly the electrostatic surface of the RIF binding pocket is dramatically changed, likely resulting in the decreased affinity for RIFs. Analysis of interactions of RMP with three common RIF mutant RNAPs suggests that modifications to RMP may recover its efficacy against RIF TB.
自1967年以来，利福平（RMP，一种利福霉素）一直被用作结核病（TB）的一线抗生素治疗药物，并且它仍然是当前短期结核病治疗的基石。耐利福霉素（RIF）结核病的发生率不断增加，其中约41%是由RNA聚合酶（RNAP）中的RpoB S531L突变引起的，这已成为全球范围内日益严重的问题。在本研究中，我们确定了含有临床上最重要的S531L突变以及另外两个常见的RIF突变体RpoB D516V和RpoB H526Y的大肠杆菌RNAP的X射线晶体结构。结构显示，在没有利福平的情况下，S531L突变对RNAP的结构或功能影响甚微（如果有影响的话）。然而，在利福平结合后，S531L突变体的利福平结合界面出现无序，这有效地降低了利福平的亲和力。相比之下，H526Y突变重塑了利福平结合口袋，产生了显著的空间冲突，基本上阻止了任何利福平的结合。虽然D516V突变体没有表现出任何此类明显的结构变化，但利福平结合口袋的静电表面肯定发生了显著变化，这可能导致对利福平的亲和力降低。对利福平与三种常见的RIF突变体RNAP相互作用的分析表明，对利福平进行修饰可能恢复其对RIF结核病的疗效。
