title: CBR antimicrobials alter coupling between the bridge helix and the β subunit in RNA polymerase.
abstract: Bacterial RNA polymerase (RNAP) is a validated target for antibacterial drugs. CBR703 series antimicrobials allosterically inhibit transcription by binding to a conserved α helix (β' bridge helix, BH) that interconnects the two largest RNAP subunits. Here we show that disruption of the BH-β subunit contacts by amino-acid substitutions invariably results in accelerated catalysis, slowed-down forward translocation and insensitivity to regulatory pauses. CBR703 partially reverses these effects in CBR-resistant RNAPs while inhibiting catalysis and promoting pausing in CBR-sensitive RNAPs. The differential response of variant RNAPs to CBR703 suggests that the inhibitor binds in a cavity walled by the BH, the β' F-loop and the β fork loop. Collectively, our data are consistent with a model in which the β subunit fine tunes RNAP elongation activities by altering the BH conformation, whereas CBRs deregulate transcription by increasing coupling between the BH and the β subunit.
authors.affiliation: Department of Biochemistry, University of Turku, Turku 20014, Finland.
authors.forename: Anssi M
authors.initials: AM
authors.lastname: Malinen
authors.affiliation: Department of Microbiology, The Ohio State University, Columbus, Ohio 43210, USA.
authors.forename: Monali
authors.initials: M
authors.lastname: Nandymazumdar
authors.affiliation: Department of Biochemistry, University of Turku, Turku 20014, Finland.
authors.forename: Matti
authors.initials: M
authors.lastname: Turtola
authors.affiliation: Department of Biochemistry, University of Turku, Turku 20014, Finland.
authors.forename: Henri
authors.initials: H
authors.lastname: Malmi
authors.affiliation: Department of Biochemistry, University of Turku, Turku 20014, Finland.
authors.forename: Thadee
authors.initials: T
authors.lastname: Grocholski
authors.affiliation: Department of Microbiology, The Ohio State University, Columbus, Ohio 43210, USA.
authors.forename: Irina
authors.initials: I
authors.lastname: Artsimovitch
authors.affiliation: Department of Biochemistry, University of Turku, Turku 20014, Finland.
authors.forename: Georgiy A
authors.initials: GA
authors.lastname: Belogurov
mesh_terms: D000578:Amidines / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D019943:Amino Acid Substitution; D000890:Anti-Infective Agents / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D055162:Biocatalysis / Q000187:drug effects; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006898:Hydroxylamines / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D000578:Amidines; D000890:Anti-Infective Agents; D001426:Bacterial Proteins; C478598:CBR 703; D006898:Hydroxylamines; D021122:Protein Subunits; D012321:DNA-Directed RNA Polymerases