Membrane efflux pumps play a major role in bacterial multidrug resistance. The tripartite multi-drug efflux pump system from Escherichia coli, AcrAB-TolC, is a target for inhibition to lessen resistance development and restore antibiotic efficacy, with homologs in other ESKAPE pathogens. Our results support a model where an inhibitor forms a molecular wedge within a cleft between the lipoyl and αβ barrel domains of AcrA, diminishing its conformational transmission of drug-evoked signals from AcrB to TolC. This work provides molecular insights into multi-drug adaptor protein function which could be valuable for developing antimicrobial therapeutics. Co-authors of the study include two School of Chemistry and Biochemistry researchers: Associate Professor James Gumbart and Ph.D. candidate Katie M. Kuo. (Gumbart is also an associate professor in the School of Physics.)