Conformational Dynamics of AcrA Govern Multidrug Efflux Pump Assembly

Abstract In Gram-negative bacteria, tripartite efflux pumps, like AcrAB-TolC from Escherichia coli, play a prominent role in the resistance against multiple antibiotics. Transport of the drugs across the outer membrane and its coupling to the electrochemical gradient is dependent on the presence of all three components. As the activity of the E. coli AcrAB-TolC efflux pump is dependent on both the concentration of substrates and the extent of the electrochemical gradient across the inner membrane, the dynamics of tripartite pump assembly and disassembly might be crucial for effective net transport of drugs towards the outside of the cell.

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Faculty Opinions recommendation of Structure of the AcrAB-TolC multidrug efflux pump.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718356663.793495679 ◽

2014 ◽

Author(s):

Paul Savage

Keyword(s):

Efflux Pump ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

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Faculty Opinions recommendation of Molecular basis for inhibition of AcrB multidrug efflux pump by novel and powerful pyranopyridine derivatives.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.726218324.793516072 ◽

2016 ◽

Author(s):

Herschel Wade

Keyword(s):

Molecular Basis ◽

Efflux Pump ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

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Faculty Opinions recommendation of Biased partitioning of the multidrug efflux pump AcrAB-TolC underlies long-lived phenotypic heterogeneity.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.727537002.793532571 ◽

2017 ◽

Author(s):

Chris Whitfield

Keyword(s):

Efflux Pump ◽

Phenotypic Heterogeneity ◽

Multidrug Efflux ◽

Multidrug Efflux Pump

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Molecular Insights into an Antibiotic Enhancer Action of New Morpholine-Containing 5-Arylideneimidazolones in the Fight against MDR Bacteria

International Journal of Molecular Sciences ◽

10.3390/ijms22042062 ◽

2021 ◽

Vol 22 (4) ◽

pp. 2062

Author(s):

Aneta Kaczor ◽

Karolina Witek ◽

Sabina Podlewska ◽

Veronique Sinou ◽

Joanna Czekajewska ◽

...

Keyword(s):

Molecular Modelling ◽

Efflux Pump ◽

Gram Negative Bacteria ◽

Potential Mechanism ◽

Multidrug Efflux ◽

Aromatic Rings ◽

Allosteric Site ◽

Multidrug Efflux Pump ◽

Dual Action

In the search for an effective strategy to overcome antimicrobial resistance, a series of new morpholine-containing 5-arylideneimidazolones differing within either the amine moiety or at position five of imidazolones was explored as potential antibiotic adjuvants against Gram-positive and Gram-negative bacteria. Compounds (7–23) were tested for oxacillin adjuvant properties in the Methicillin-susceptible S. aureus (MSSA) strain ATCC 25923 and Methicillin-resistant S. aureus MRSA 19449. Compounds 14–16 were tested additionally in combination with various antibiotics. Molecular modelling was performed to assess potential mechanism of action. Microdilution and real-time efflux (RTE) assays were carried out in strains of K. aerogenes to determine the potential of compounds 7–23 to block the multidrug efflux pump AcrAB-TolC. Drug-like properties were determined experimentally. Two compounds (10, 15) containing non-condensed aromatic rings, significantly reduced oxacillin MICs in MRSA 19449, while 15 additionally enhanced the effectiveness of ampicillin. Results of molecular modelling confirmed the interaction with the allosteric site of PBP2a as a probable MDR-reversing mechanism. In RTE, the compounds inhibited AcrAB-TolC even to 90% (19). The 4-phenylbenzylidene derivative (15) demonstrated significant MDR-reversal “dual action” for β-lactam antibiotics in MRSA and inhibited AcrAB-TolC in K. aerogenes. 15 displayed also satisfied solubility and safety towards CYP3A4 in vitro.

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Insight into the AcrAB-TolC Complex Assembly Process Learned from Competition Studies

Antibiotics ◽

10.3390/antibiotics10070830 ◽

2021 ◽

Vol 10 (7) ◽

pp. 830

Author(s):

Prasangi Rajapaksha ◽

Isoiza Ojo ◽

Ling Yang ◽

Ankit Pandeya ◽

Thilini Abeywansha ◽

...

Keyword(s):

Efflux Pump ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Assembly Process ◽

Multidrug Efflux ◽

E Coli ◽

Multidrug Efflux Pumps ◽

Negative Effect ◽

Dynamic Assembly ◽

Pump Assembly

The RND family efflux pump AcrAB-TolC in E. coli and its homologs in other Gram-negative bacteria are major players in conferring multidrug resistance to the cells. While the structure of the pump complex has been elucidated with ever-increasing resolution through crystallography and Cryo-EM efforts, the dynamic assembly process remains poorly understood. Here, we tested the effect of overexpressing functionally defective pump components in wild type E. coli cells to probe the pump assembly process. Incorporation of a defective component is expected to reduce the efflux efficiency of the complex, leading to the so called “dominant negative” effect. Being one of the most intensively studied bacterial multidrug efflux pumps, many AcrA and AcrB mutations have been reported that disrupt efflux through different mechanisms. We examined five groups of AcrB and AcrA mutants, defective in different aspects of assembly and substrate efflux. We found that none of them demonstrated the expected dominant negative effect, even when expressed at concentrations many folds higher than their genomic counterpart. The assembly of the AcrAB-TolC complex appears to have a proof-read mechanism that effectively eliminated the formation of futile pump complex.

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Correlation between the Antibiotic Resistance Genes and Susceptibility to Antibiotics among the Carbapenem-Resistant Gram-Negative Pathogens

Antibiotics ◽

10.3390/antibiotics10030255 ◽

2021 ◽

Vol 10 (3) ◽

pp. 255

Author(s):

Salma M. Abdelaziz ◽

Khaled M. Aboshanab ◽

Ibrahim S. Yahia ◽

Mahmoud A. Yassien ◽

Nadia A. Hassouna

Keyword(s):

Antibiotic Resistance ◽

Resistance Genes ◽

Efflux Pump ◽

Antibiotic Resistance Genes ◽

Multiple Drug ◽

P Value ◽

Multidrug Efflux ◽

Multidrug Efflux Pump ◽

Gram Negative ◽

Carbapenem Resistant

In this study, the correlation between the antibiotic resistance genes and antibiotic susceptibility among the carbapenem-resistant Gram-negative pathogens (CRGNPs) recovered from patients diagnosed with acute pneumonia in Egypt was found. A total of 194 isolates including Klebsiella pneumoniae (89; 46%), Escherichia coli (47; 24%) and Pseudomonas aeruginosa (58; 30%) were recovered. Of these, 34 (18%) isolates were multiple drug resistant (MDR) and carbapenem resistant. For the K. pneumoniae MDR isolates (n = 22), blaNDM (14; 64%) was the most prevalent carbapenemase, followed by blaOXA-48 (11; 50%) and blaVIM (4; 18%). A significant association (p value < 0.05) was observed between the multidrug efflux pump (AcrA) and resistance to β-lactams and the aminoglycoside acetyl transferase gene (aac-6’-Ib) gene and resistance to ciprofloxacin, azithromycin and β-lactams (except for aztreonam). For P. aeruginosa, a significant association was noticed between the presence of the blaSHV gene and the multidrug efflux pump (MexA) and resistance to fluoroquinolones, amikacin, tobramycin, co-trimoxazole and β-lactams and between the aac-6’-Ib gene and resistance to aminoglycosides. All P. aeruginosa isolates (100%) harbored the MexAB-OprM multidrug efflux pump while 86% of the K. pneumoniae isolates harbored the AcrAB-TolC pump. Our results are of great medical importance for the guidance of healthcare practitioners for effective antibiotic prescription.

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