Ligand-transporter interaction in the AcrB multidrug efflux pump determined by fluorescence polarization assay

The AcrB of Escherichia coli   pumps out a wide range of compounds, including most of the currently available antibiotics, and contributes significantly to the serious problem of multidrug resistance of pathogenic bacteria. Quantitative analysis of drug efflux by this pump requires the measurement of the affinity of ligands. Yet there has been no success in determining these values. We introduce here an approach of steady-state fluorescence polarization to study the interactions between four different ligands and the purified AcrB transporter in a detergent environment. Our assays indicate that the transporter binds these drugs with KDKD values ranging from 5.5 to 74.1 μM.