Disassembly of the Escherichia coli AcrABZ-TolC efflux pump by ligand-mediated disruption of TolC-AcrA interfacial contacts

Szal, T.; Petsolari, E.; Veliks, J.; Cruz, C. D.; Paunina, L.; Madre, M.; Rachad, F.-Z.; Lewe, P.; Witt, S.; Tammela, P.; Jirgensons, A.; Luisi, B. F.; Windshügel, B.

doi:10.1101/2025.02.18.638599

bioRxiv · 10.1101/2025.02.18.638599

Disassembly of the Escherichia coli AcrABZ-TolC efflux pump by ligand-mediated disruption of TolC-AcrA interfacial contacts

Szal, T.·Petsolari, E.·Veliks, J.·Cruz, C. D.·Paunina, L.·Madre, M.·Rachad, F.-Z.·Lewe, P.·Witt, S.·Tammela, P.·Jirgensons, A.·Luisi, B. F.·Windshügel, B.

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Abstract

The outer membrane factor TolC is an essential component of various efflux pump complexes in E. coli and represents a potential target for antibiotic adjuvants. By means of a virtual screen for TolC-binding compounds, we identified the kinase inhibitor CEP- 37440 to shift the minimum inhibitory concentration of antibiotics piperacillin and levofloxacin in E. coli. To determine the substructure relevant for TolC binding, a hit deconstruction approach was applied, resulting in a fragment-like compound with low affinity for TolC and AcrB (LP-115). Dynamic light scattering revealed LP-115 to reduce the hydrodynamic radius of AcrABZ-TolC, indicating a disassembly of the efflux pump complex. A cryo-EM structure demonstrated LP-115 to bind at the TolC-AcrA interface within the AcrABZ-TolC complex, thereby disordering the interface and inducing a closed conformation of TolC. Our results suggest that ligand-mediated TolC-AcrA interface disruption represents a novel mechanism of efflux pump inhibition.

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Szal, T., Petsolari, E., Veliks, J., Cruz, C. D., Paunina, L., Madre, M., Rachad, F.-Z., Lewe, P., Witt, S., Tammela, P., Jirgensons, A., Luisi, B. F., Windshügel, B.. 2025-02-19. Disassembly of the Escherichia coli AcrABZ-TolC efflux pump by ligand-mediated disruption of TolC-AcrA interfacial contacts. https://doi.org/10.1101/2025.02.18.638599

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