Substrate-bound and substrate-free outward-facing structures of a multidrug ABC exporter

Area of Science:

• Structural Biology
• Biochemistry
• Molecular Biology

Background:

• Multidrug ABC transporters are crucial for cellular drug efflux.
• The precise mechanism of drug release from these transporters remains poorly understood.
• BmrA from Bacillus subtilis is a well-characterized multidrug ABC transporter.

Purpose of the Study:

• To elucidate the molecular mechanism of drug release in multidrug ABC transporters.
• To determine the structure of BmrA in complex with its substrate rhodamine 6G (R6G).
• To investigate the conformational changes associated with substrate binding and release.

Main Methods:

• X-ray crystallography to resolve high-resolution structures.
• Single-particle cryo-electron microscopy (cryo-EM) in detergent solution.
• Hydrogen/deuterium exchange and molecular dynamics simulations to assess flexibility and dynamics.

Main Results:

• Three ATP-Mg2+-bound outward-facing conformations of BmrA were resolved.
• A structure captured two R6G molecules within the drug-binding cavity.
• R6G binding induces rearrangements in transmembrane helices, indicating local flexibility.
• Simulations revealed R6G's role in maintaining the cavity's openness post-release.

Conclusions:

• The study provides unprecedented structural insights into the drug release mechanism of BmrA.
• Substrate interaction with transmembrane helices is key to modulating transporter conformation.
• Understanding these mechanisms can inform the development of strategies to overcome multidrug resistance.