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Multidrug/dye/detergent/bile salt/organic solvent resistance pump (substrates include: chloramphenicol, tetracycline, erythromycin, nalidixic acid, fusidic acid, fluoroquinolones, lipophilic β-lactams, norfloxacin, doxorubicin, novobiocin, rifampin, trimethoprim, acriflavin, crystal violet, ethidium, disinfectants, rhodamine-6G, TPP, benzalkonium, SDS, Triton X-100, deoxycholate/bile salts/organic solvents (alkanes), growth inhibitory steroid hormones (estradiola and progesterone), and phospholipids) (Elkins and Mullis, 2006). Lateral entry of substrates from the lipid bilayer into AcrB and its homologues has been proposed (Yu et al., 2003a; 2003b). [An asymmetric trimeric structure is established with AcrA having a hexameric structure, and TolC having a trimeric structure (Seeger et al., 2006]. A structure of a complex with YajC is also known (Törnroth-Horsefield et al., 2007). A covalently linked trimer of AcrB provides evidence for a peristaltic pump, alternative access, rotation mechanism (Takatsuka and Nikaido, 2009;Nikaido and Takatsuka, 2009; Pos, 2009) Further evidence for a rotatory mechanisms stems from kinetic analyses for cephalosporin efflux which can exhibit positive cooperativity (Nagano and Nikaido, 2009). May also export signaling molecules for cell-cell communication (Yang et al., 2006). The substrates may be captured in the lower cleft region of AcrB, then transported through the binding pocket, the gate, and finally to the AcrA funnel that connects AcrB to TolC (Husain & Nikaido et al., 2010).  AcrB has been converted into a light-driven proton pump using delta-rhodopsin (dR) linked to AcrB via a glycophorin A transmembrane domain. This created a solar powered protein capable of selectively capturing antibiotics from bulk solutions (Kapoor and Wendell 2013).  The trimeric structure is essential for activity (Ye et al. 2014).  Association with AcrZ (TC# 8.A.50), a small 1 TMS protein (49 aas) that modifies the substrate specificity of AcrAB, has been demonstrated (Hobbs et al. 2012).  In a similar way, the binding of YajC to AcrB stimulates the export of ampicillin (Törnroth-Horsefield et al. 2007). AcrZ binds to AcrB in a concave surface of the transmembrane domain (Du et al. 2015).  Substrate binding accelerates conformational transitions and substrate dissociation, demonstrating cooperativity (Wang et al. 2015). The overall structure of AcrAB-TolC exemplifies the adaptor bridging model, wherein the funnel-like AcrA hexamer forms an intermeshing cogwheel interaction with the alpha-barrel tip region of TolC. Direct interaction between AcrB and TolC is not allowed (Kim et al. 2015).  TMS2 in AcrB is required for lipophilic carboxylate binding. A groove shaped by the interface between TMS1 and TMS2 specifically binds fusidic acid and other lipophilic carboxylated drugs (Oswald et al. 2016). After ligand binding, a proton may bind to an acidic residue(s) in the transmembrane domain, i.e., Asp407 or Asp408, within the putative network of electrostatically interacting residues, which also include Lys940 and Thr978, and this may initiate a series of conformational changes that result in drug expulsion (Su et al. 2006). His978 is probably on the H+ pathway (Takatsuka and Nikaido 2006). AcrAB-TolC segregates to the old pole following cell division, causing the two daughter cells to exhibit different drug resistances (Bergmiller et al. 2017). The hoisting-loop is a highly flexible hinge that enables conformational energy transmission (Zwama et al. 2017). AcrB exhibits three distinct conformational states in the transport cycle, substrate access, binding, and extrusion, or loose (L), tight (T), and open (O) states, respectively (Yue et al. 2017). Simulations show that both Asp407 and Asp408 are deprotonated in the L/T states, while only Asp408 is protonated in the O state. Release of a proton from Asp408 in the O state results in large conformational changes.  Simulations offer dynamic details of how proton release drives the O-to-L transition in AcrB (Yue et al. 2017).  The three-dimensional structures of the homo-trimer complexes of AcrB-like transporters, and a three-step functional rotation helps to explain the mechanism of transport, but a more comprehensive model has been proposed (Zhang et al. 2017). Preparation of the trimeric complex (AcrAB/TolC) for cryo EM has been described (Du et al. 2018). The structural and energetic basis behind coupling functional rotation to proton translocation has been presented (Matsunaga et al. 2018). Protonation of  transmembrane Asp408 in the drug-bound protomer drives rotation. The conformational pathway identifies vertical shear motions among several transmembrane helices, which regulate alternate access of water as well as peristaltic motions that pump drugs into the periplasm (Matsunaga et al. 2018). CryoEM of detergent-free AcrB preserves lipid-protein interactions for visualization and reveals how the lipids pack against the protein (Qiu et al. 2018). In the presence of translation-inhibiting antibiotics, resistance acquisition depends on the AcrAB-TolC multidrug efflux pump, because it reduces tetracycline concentrations in the cell. Protein synthesis can thus persist and TetA expression can be initiated immediately after plasmid acquisition. AcrAB-TolC efflux activity can also preserve resistance acquisition by plasmid transfer in the presence of antibiotics with other modes of action (Nolivos et al. 2019). Multiple transport pathways within AcrB are tuned to substrate physicochemical properties related to the polyspecificity of the pump (Tam et al. 2019). The cryoEM structure of AcrB in an artificial membrane at 3.9 Å resolution has been solved (Yao et al. 2020). The indole-dependent transport mechanism has been examined (Jewel et al. 2020). AcrB subunits rimerize to form the minimal functional unit, stabilized noncovalently by helix-helix interactions between TMS 1 and TMS 8. Peptides resembling these two TMSs inhibit subunit associations and drug export activity (Jesin et al. 2020). AcrB cycles between three functionally interdependent protomers through the loose (L), tight (T) and open (O) states during cooperative catalysis. Tam et al. 2021 presented 13 X-ray structures of AcrB in intermediate states of the transport cycle. Structure-based mutational analysis combined with drug susceptibility assays indicated 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 showed binding of fusidic acid deeply inside the T protomer transmembrane domain. Thiol cross-link substrate protection assays indicated 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 the presence of multiple drugs (Tam et al. 2021).

Accession Number:P0AAW9
Protein Name:Multidrug efflux pump accessory protein AcrZ
Molecular Weight:5300.00
Species:Escherichia coli (strain K12) [83333]
Number of TMSs:1
Location1 / Topology2 / Orientation3: Cell inner membrane1 / Single-pass membrane protein2
Substrate phospholipid, steroid hormone, bile acid, ethidium, disinfectant, drug, dye, detergent

Cross database links:

4C48   5NG5     

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