In situ structure of the AcrAB-TolC efflux pump at subnanometer resolution

• Published in: Structure (London) 2022-01, Vol.30 (1), p.107-113.e3
• Main Authors: Chen, Muyuan, Shi, Xiaodong, Yu, Zhili, Fan, Guizhen, Serysheva, Irina I., Baker, Matthew L., Luisi, Ben F., Ludtke, Steven J., Wang, Zhao
• Format: Article
• Language: English
• Subjects: AcrAB-TolC; Allosteric Regulation; Bacterial Outer Membrane Proteins - chemistry; Bacterial Outer Membrane Proteins - metabolism; Carrier Proteins - chemistry; cryo-ET; Cryoelectron Microscopy; Electron Microscope Tomography; Escherichia coli - chemistry; Escherichia coli - metabolism; Escherichia coli Proteins - chemistry; Escherichia coli Proteins - metabolism; in situ; Ligands; Lipoproteins - chemistry; Lipoproteins - metabolism; Membrane Transport Proteins - chemistry; Membrane Transport Proteins - metabolism; Models, Molecular; multidrug efflux pump; Multidrug Resistance-Associated Proteins - chemistry; Multidrug Resistance-Associated Proteins - metabolism; Protein Conformation; subnanometer resolution
• ISSN: 0969-2126; 1878-4186
• DOI: 10.1016/j.str.2021.08.008

The tripartite AcrAB-TolC assembly, which spans both the inner and outer membranes in Gram-negative bacteria, is an efflux pump that contributes to multidrug resistance. Here, we present the in situ structure of full-length Escherichia coli AcrAB-TolC determined at 7 Å resolution by electron cryo-tomography. The TolC channel penetrates the outer membrane bilayer through to the outer leaflet and exhibits two different configurations that differ by a 60° rotation relative to the AcrB position in the pump assembly. AcrA protomers interact directly with the inner membrane and with AcrB via an interface located in proximity to the AcrB ligand-binding pocket. Our structural analysis suggests that these AcrA-bridged interactions underlie an allosteric mechanism for transmitting drug-evoked signals from AcrB to the TolC channel within the pump. Our study demonstrates the power of in situ electron cryo-tomography, which permits critical insights into the function of bacterial efflux pumps. [Display omitted] •The in situ structure of AcrAB-TolC efflux pump is achieved at 7 Å resolution•The TolC channel penetrates the outer membrane bilayer through to the outer leaflet•Domain interactions underlie an allosteric mechanism for pump opening and closing•Cellular cryo-ET reveals critical structural details relevant to pump activity The AcrAB-TolC assembly is a key efflux pump that contributes to multidrug resistance within Gram-negative bacteria. Spanning the cell envelope, it is a challenging target for in vitro structural studies. Chen et al. solve the subnanometer in situ structure of AcrAB-TolC, suggesting an allosteric mechanism during drug efflux.