title: Structural basis for rifamycin resistance of bacterial RNA polymerase by the three most clinically important RpoB mutations found in Mycobacterium tuberculosis.
abstract: 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 (RIFR) 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 RIFRmutants, 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 RIFRmutant RNAPs suggests that modifications to RMP may recover its efficacy against RIFRTB.
authors.affiliation: Department of Biochemistry and Molecular Biology, The Center for RNA Molecular Biology, The Pennsylvania State University, University Park, PA, 16802, USA.
authors.forename: Vadim
authors.initials: V
authors.lastname: Molodtsov
authors.affiliation: Department of Medicinal Chemistry, College of Pharmacy, University of Michigan, 428 Church St, Ann Arbor, MI, 48109-1065, USA.
authors.forename: Nathan T
authors.initials: NT
authors.lastname: Scharf
authors.affiliation: Department of Medicinal Chemistry, College of Pharmacy, University of Michigan, 428 Church St, Ann Arbor, MI, 48109-1065, USA.
authors.forename: Maxwell A
authors.initials: MA
authors.lastname: Stefan
authors.affiliation: Department of Medicinal Chemistry, College of Pharmacy, University of Michigan, 428 Church St, Ann Arbor, MI, 48109-1065, USA.
authors.forename: George A
authors.initials: GA
authors.lastname: Garcia
authors.affiliation: Department of Biochemistry and Molecular Biology, The Center for RNA Molecular Biology, The Pennsylvania State University, University Park, PA, 16802, USA.
authors.forename: Katsuhiko S
authors.identifier: 0000-0003-2244-0501
authors.initials: KS
authors.lastname: Murakami
mesh_terms: D000904:Antibiotics, Antitubercular / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics*; D001665:Binding Sites / Q000235:genetics; D018360:Crystallography, X-Ray; D012321:DNA-Directed RNA Polymerases / Q000187:drug effects / Q000235:genetics / Q000648:ultrastructure*; D024881:Drug Resistance, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D009154:Mutation / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics; D011487:Protein Conformation; D012329:RNA, Bacterial; D012293:Rifampin / Q000494:pharmacology*; D014397:Tuberculosis, Pulmonary / Q000188:drug therapy* / Q000382:microbiology
chemical_list: D000904:Antibiotics, Antitubercular; D001426:Bacterial Proteins; D012329:RNA, Bacterial; C505805:rpoB protein, Mycobacterium tuberculosis; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin