Functional relevance of AcrB Trimerization in pump assembly and substrate binding

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• AcrB is a key multidrug transporter in Escherichia coli's inner membrane.
• It functions as an obligate homotrimer, forming an efflux complex with AcrA and TolC.
• AcrB's function relies on proton translocation, substrate binding/transport, interaction with AcrA/TolC, and homotrimer formation.

Purpose of the Study:

• To investigate the interdependence of AcrB trimerization and AcrA interaction.
• To examine how protein-protein interactions affect substrate binding and transport in AcrB.

Main Methods:

• Chemical crosslinking was used to study AcrA-AcrB interactions.
• In vivo fluorescent labeling was employed to probe substrate binding dynamics.
• Functional assays assessed the impact of trimerization and proton influx on substrate binding.

Main Results:

• Dissociation of the AcrB homotrimer significantly reduced its interaction with AcrA.
• Substrate binding to AcrB was independent of the presence of AcrA and TolC.
• AcrB trimerization and proton influx capability influenced substrate binding at specific sites.

Conclusions:

• AcrB trimerization is crucial for its interaction with AcrA.
• While AcrA and TolC are not essential for initial substrate binding to AcrB, AcrB's functional state (trimerization, proton translocation) impacts substrate interaction within the transporter.