title: Unconventional surface plasmon resonance signals reveal quantitative inhibition of transcriptional repressor EthR by synthetic ligands.
abstract: EthR is a mycobacterial repressor that limits the bioactivation of ethionamide, a commonly used anti-tuberculosis second-line drug. Several efforts have been deployed to identify EthR inhibitors abolishing the DNA-binding activity of the repressor. This led to the demonstration that stimulating the bioactivation of Eth through EthR inhibition could be an alternative way to fight Mycobacterium tuberculosis. We propose a new surface plasmon resonance (SPR) methodology to study the affinity between inhibitors and EthR. Interestingly, the binding between inhibitors and immobilized EthR produced a dose-dependent negative SPR signal. We demonstrate that this signal reveals the affinity of small molecules for the repressor. The affinity constants (K(D)) correlate with their capacity to inhibit the binding of EthR to DNA. We hypothesize that conformational changes in EthR during ligand interaction could be responsible for this SPR signal. Practically, this unconventional result opens perspectives onto the development of an SPR assay that would at the same time reveal structural changes in the target upon binding with an inhibitor and the binding constant of this interaction.
authors.affiliation: Biostructures and Drug Discovery, INSERM U761, F-59000 Lille, France; Faculté de Pharmacie de Lille, University of Lille Nord de France, F-59000 Lille, France; Institut Pasteur de Lille, F-59019 Lille, France; IFR 142, Molecular and Cellular Medicine, F-59021 Lille, France; PRIM, F-59019 Lille, France.
authors.forename: Céline
authors.initials: C
authors.lastname: Crauste
authors.affiliation: Biostructures and Drug Discovery, INSERM U761, F-59000 Lille, France; Faculté de Pharmacie de Lille, University of Lille Nord de France, F-59000 Lille, France; Institut Pasteur de Lille, F-59019 Lille, France; IFR 142, Molecular and Cellular Medicine, F-59021 Lille, France; PRIM, F-59019 Lille, France. Electronic address: nicolas.willand@univ-lille2.fr.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Willand
authors.affiliation: Biostructures and Drug Discovery, INSERM U761, F-59000 Lille, France; Faculté de Pharmacie de Lille, University of Lille Nord de France, F-59000 Lille, France; Institut Pasteur de Lille, F-59019 Lille, France; IFR 142, Molecular and Cellular Medicine, F-59021 Lille, France; PRIM, F-59019 Lille, France.
authors.forename: Baptiste
authors.initials: B
authors.lastname: Villemagne
authors.affiliation: Biostructures and Drug Discovery, INSERM U761, F-59000 Lille, France; Faculté de Pharmacie de Lille, University of Lille Nord de France, F-59000 Lille, France; Institut Pasteur de Lille, F-59019 Lille, France; IFR 142, Molecular and Cellular Medicine, F-59021 Lille, France; PRIM, F-59019 Lille, France.
authors.forename: Marion
authors.initials: M
authors.lastname: Flipo
authors.affiliation: Institut Pasteur de Lille, F-59019 Lille, France; PRIM, F-59019 Lille, France; Center for Infection and Immunity of Lille, INSERM U1019-CNRS UMR 8204, University of Lille Nord de France, F-59000 Lille, France.
authors.forename: Eve
authors.initials: E
authors.lastname: Willery
authors.affiliation: Institut Pasteur de Lille, F-59019 Lille, France; PRIM, F-59019 Lille, France; Center for Infection and Immunity of Lille, INSERM U1019-CNRS UMR 8204, University of Lille Nord de France, F-59000 Lille, France.
authors.forename: Xavier
authors.initials: X
authors.lastname: Carette
authors.affiliation: Institut Pasteur de Lille, F-59019 Lille, France; PRIM, F-59019 Lille, France; Center for Infection and Immunity of Lille, INSERM U1019-CNRS UMR 8204, University of Lille Nord de France, F-59000 Lille, France.
authors.forename: Martin Moune
authors.initials: MM
authors.lastname: Dimala
authors.affiliation: Molecular Interactions Platform, Institute of Predictive Medicine and Therapeutic Research, IFR 114, Faculty of Medicine, Lille 2 University of Health and Law, F-59045 Lille, France.
authors.forename: Anne-Sophie
authors.initials: AS
authors.lastname: Drucbert
authors.affiliation: Molecular Interactions Platform, Institute of Predictive Medicine and Therapeutic Research, IFR 114, Faculty of Medicine, Lille 2 University of Health and Law, F-59045 Lille, France.
authors.forename: Pierre-Marie
authors.initials: PM
authors.lastname: Danze
authors.affiliation: Biostructures and Drug Discovery, INSERM U761, F-59000 Lille, France; Faculté de Pharmacie de Lille, University of Lille Nord de France, F-59000 Lille, France; Institut Pasteur de Lille, F-59019 Lille, France; IFR 142, Molecular and Cellular Medicine, F-59021 Lille, France; PRIM, F-59019 Lille, France.
authors.forename: Benoit
authors.initials: B
authors.lastname: Deprez
authors.affiliation: Institut Pasteur de Lille, F-59019 Lille, France; PRIM, F-59019 Lille, France; Center for Infection and Immunity of Lille, INSERM U1019-CNRS UMR 8204, University of Lille Nord de France, F-59000 Lille, France. Electronic address: alain.baulard@pasteur-lille.fr.
authors.forename: Alain R
authors.initials: AR
authors.lastname: Baulard
mesh_terms: D019921:Biotinylation; D008024:Ligands; D009169:Mycobacterium tuberculosis; D012097:Repressor Proteins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism*; D020349:Surface Plasmon Resonance / Q000379:methods*; D044366:Transition Temperature
chemical_list: C481661:EthR protein, Mycobacterium tuberculosis; D008024:Ligands; D012097:Repressor Proteins
keywords: EthR; SPR negative signal; Surface plasmon resonance; TetR; Transcriptional regulator; Tuberculosis