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bioRxiv · 10.1101/2025.10.29.685381

Structural mechanisms of pump assembly and drug transport in the AcrAB-TolC efflux system

Ge, X.·Gu, Z.·Wang, J.
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Abstract

Tripartite multidrug efflux pumps that span the cell envelope are essential for antibiotic resistance in Gram-negative bacteria. Here, we report cryo-EM structures of two endogenous efflux complexes from Escherichia coli: a TolC-YbjP subcomplex at 3.56 [A] resolution and the complete TolC-YbjP-AcrABZ pump at 3.39 [A]. Structural analysis reveals that YbjP, a previously uncharacterized lipoprotein, binds TolC in a 3:3 stoichiometry, bridging the TolC protomers at their equatorial domain. Clear density of the mature YbjPs N-terminal Cys19 indicates that YbjP is anchored to the outer membrane by an N-terminal lipid moiety. Notably, YbjP remains bound as TolC undergoes AcrA-induced opening, suggesting that this accessory protein accommodates the conformational change. The AcrB trimer simultaneously presents three distinct conformational states (L, T, O), capturing a complete transport cycle. These high-resolution structures provide insights into the architecture and mechanism of clinically relevant efflux machinery, identifying YbjP as a previously unrecognized structural component that contributes to TolC positioning and may assist in its membrane localization.

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Ge, X., Gu, Z., Wang, J.. 2025-10-30. Structural mechanisms of pump assembly and drug transport in the AcrAB-TolC efflux system. https://doi.org/10.1101/2025.10.29.685381

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