title: Molecular insights into the mechanism of phenotypic tolerance to rifampicin conferred on mycobacterial RNA polymerase by MsRbpA.
abstract: The protein MsRbpA from Mycobacterium smegmatis rescues RNA polymerase (RNAP) from the inhibitory effect of rifampicin (Rif). We have reported previously that MsRbpA interacts with the β-subunit of RNAP and that the effect of MsRbpA on Rif-resistant (Rif(R)) RNAP is minimal. Here we attempted to gain molecular insights into the mechanism of action of this protein with respect to its role in rescuing RNAP from Rif-mediated transcription inhibition. Our experimental approach comprised multiple-round transcription assays, fluorescence spectroscopy, MS and surface plasmon resonance in order to meet the above objective. Based on our molecular studies we propose here that Rif is released from its binding site in the RNAP-Rif complex in the presence of MsRbpA. Biophysical studies reveal that the location of MsRbpA on RNAP is at the junction of the β- and β'-subunits, close to the Rif-binding site and the (i+1) site on RNAP.
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Abhinav
authors.initials: A
authors.lastname: Dey
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Amit Kumar
authors.initials: AK
authors.lastname: Verma
authors.affiliation: Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.
authors.forename: Dipankar
authors.initials: D
authors.lastname: Chatterji
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000187:drug effects / Q000378:metabolism; D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013058:Mass Spectrometry; D020102:Mycobacterium smegmatis / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000378:metabolism; D012293:Rifampin / Q000378:metabolism / Q000494:pharmacology*; D016415:Sequence Alignment; D013050:Spectrometry, Fluorescence; D020349:Surface Plasmon Resonance; D014158:Transcription, Genetic / Q000187:drug effects*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D021122:Protein Subunits; D016601:RNA-Binding Proteins; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin