Rhodamine6G and Hœchst33342 narrow BmrA conformational spectrum for a more efficient use of ATP
Multidrug ABC transporters harness the energy of ATP binding and hydrolysis to translocate substrates out of the cell and detoxify them. While this involves a well-accepted alternating access mechanism, molecular details of this interplay are still elusive. Rhodamine6G binding on a catalytic inactive mutant of the homodimeric multidrug ABC transporter BmrA triggers a cooperative binding of ATP on the two identical nucleotide-binding-sites, otherwise michaelian. Here, we investigate this asymmetric behavior via a structural-enzymology approach, solving cryoEM structures of BmrA at defined ATP ratios, highlighting the plasticity of BmrA as it undergoes the transition from inward to outward facing conformations. Analysis of continuous heterogeneity within cryoEM data and structural dynamics, reveals that Rhodamine6G narrows the conformational spectrum explored by the nucleotide-binding domains. We observe the same behavior for the other drug Hœchst33342. Following on these findings, the effect of drug-binding showed an ATPase stimulation and a maximal transport activity of the wild-type protein at the concentration-range where the cooperative transition occurs. Altogether, these findings provide a description of the influence of drug binding on the ATP-binding sites through a change in conformational dynamics.
Authors
- Alexia Gobet (ORCID: https://orcid.org/0000-0002-8294-8633)
- Cédric Orelle (ORCID: https://orcid.org/0000-0003-3418-3290)
- Sandrine Magnard (ORCID: https://orcid.org/0000-0003-1195-1840)
- Juliette Martin (ORCID: https://orcid.org/0000-0002-4787-0885)
- Loïck Moissonnier (ORCID: https://orcid.org/0000-0002-7223-9454)
- Vincent Chaptal (ORCID: https://orcid.org/0000-0003-2603-0294)
- Jean‐Michel Jault (ORCID: https://orcid.org/0000-0003-1743-2777)
- Guy Schoehn (ORCID: https://orcid.org/0000-0002-1459-3201)
- Pierre Falson (ORCID: https://orcid.org/0000-0002-9760-4577)
- Eleftherios Zarkadas (ORCID: https://orcid.org/0000-0002-9276-9831)
- E. Bettler (ORCID: https://orcid.org/0000-0001-8400-8277)
- Raphaël Terreux (ORCID: https://orcid.org/0000-0003-1243-7413)
Institutions
- Université Claude Bernard Lyon 1 (FR)
- École Normale Supérieure de Lyon (FR)
- Centre National de la Recherche Scientifique (FR)
- Inserm (FR)
- Aarhus University (DK)
- Commissariat à l'Énergie Atomique et aux Énergies Alternatives (FR)
- CEA Grenoble (FR)
- Structural and Molecular Basis of Infectious Systems (FR)
- Institut de Biologie Structurale (FR)
- Laboratoire de Biologie Tissulaire et d'Ingénierie Thérapeutique (FR)
- Université Grenoble Alpes (FR)
Publication Details
- Journal
- Nature Communications
- Published
- 2025-02-18
- DOI
- https://doi.org/10.1038/s41467-025-56849-z
- Primary Topic
- Drug Transport and Resistance Mechanisms
- Type
- article
- Field-Weighted Citation Impact
- 0.00
Funders
- Infrastructures en Biologie Santé et Agronomie
- Agence Nationale de la Recherche
- European Synchrotron Radiation Facility
- Fondation pour la Recherche Médicale
- Centre National de la Recherche Scientifique
- Région Auvergne-Rhône-Alpes
- French Infrastructure for Integrated Structural Biology
- Institut National de Physique Nucléaire et de Physique des Particules