Allosteric drug transport mechanism of multidrug transporter AcrB

Gram-negative bacteria maintain an intrinsic resistance mechanism against entry of noxious compounds by utilizing highly efficient efflux pumps. The E. coli AcrAB-TolC drug efflux pump contains the inner membrane H+/drug antiporter AcrB comprising three functionally interdependent protomers, cycling consecutively through the loose (L), tight (T) and open (O) state during cooperative catalysis. Here, we present 13 X-ray structures of AcrB in intermediate states of the transport cycle. Structure-based mutational analysis combined with drug susceptibility assays indicate that drugs are guided through dedicated transport channels toward the drug binding pockets. A co-structure obtained in the combined presence of erythromycin, linezolid, oxacillin and fusidic acid shows binding of fusidic acid deeply inside the T protomer transmembrane domain. Thiol cross-link substrate protection assays indicate that this transmembrane domain-binding site can also accommodate oxacillin or novobiocin but not erythromycin or linezolid. AcrB-mediated drug transport is suggested to be allosterically modulated in presence of multiple drugs. Gram-negative bacteria can display intrinsic antibiotic resistance due to the action of tripartite efflux pumps, which include a H+/drug antiporter component. Here, the authors present a structure-function analysis of antiporter AcrB in intermediate states of the transport cycle, showing novel drug-binding sites and transport pathways.

Automated summary

Gram-negative bacteria maintain intrinsic resistance against noxious compounds through highly efficient efflux pumps. The AcrB drug efflux pump of E. coli, comprising three interdependent protomers, cycles through different states during cooperative catalysis. Structural analysis reveals novel drug-binding sites and transport pathways, suggesting allosteric modulation of drug transport in the presence of multiple drugs.