Computational Study of Correlated Domain Motions in the AcrB Efflux Transporter

As active part of the major efflux system inE. colibacteria, AcrB is responsible for the uptake and pumping of toxic substrates from the periplasm toward the extracellular space. In combination with the channel protein TolC and membrane fusion protein AcrA, this efflux pump is able to help the bacterium to survive different kinds of noxious compounds. With the present study we intend to enhance the understanding of the interactions between the domains and monomers, for example, the transduction of mechanical energy from the transmembrane domain into the porter domain, correlated motions of different subdomains within monomers, and cooperative effects between monomers. To this end, targeted molecular dynamics simulations have been employed either steering the whole protein complex or specific parts thereof. By forcing only parts of the complex towards specific conformational states, the risk for transient artificial conformations during the simulations is reduced. Distinct cooperative effects between the monomers in AcrB have been observed. Possible allosteric couplings have been identified providing microscopic insights that might be exploited to design more efficient inhibitors of efflux systems.

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Pyridylpiperazine-based allosteric inhibitors of RND-type multidrug efflux pumps

Nature Communications ◽

10.1038/s41467-021-27726-2 ◽

2022 ◽

Vol 13 (1) ◽

Author(s):

Coline Plé ◽

Heng-Keat Tam ◽

Anais Vieira Da Cruz ◽

Nina Compagne ◽

Juan-Carlos Jiménez-Castellanos ◽

...

Keyword(s):

Transmembrane Domain ◽

Efflux Pump ◽

Binding Pocket ◽

Multidrug Efflux ◽

Allosteric Inhibitors ◽

E Coli ◽

Multidrug Efflux Pumps ◽

Proton Relay ◽

Rnd Transporter ◽

Dynamics Simulations

AbstractEfflux transporters of the RND family confer resistance to multiple antibiotics in Gram-negative bacteria. Here, we identify and chemically optimize pyridylpiperazine-based compounds that potentiate antibiotic activity in E. coli through inhibition of its primary RND transporter, AcrAB-TolC. Characterisation of resistant E. coli mutants and structural biology analyses indicate that the compounds bind to a unique site on the transmembrane domain of the AcrB L protomer, lined by key catalytic residues involved in proton relay. Molecular dynamics simulations suggest that the inhibitors access this binding pocket from the cytoplasm via a channel exclusively present in the AcrB L protomer. Thus, our work unveils a class of allosteric efflux-pump inhibitors that likely act by preventing the functional catalytic cycle of the RND pump.

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A Model for Allosteric Communication in Drug Transport by the AcrAB-TolC Tripartite Efflux Pump

Antibiotics ◽

10.3390/antibiotics11010052 ◽

2022 ◽

Vol 11 (1) ◽

pp. 52

Author(s):

Anya Webber ◽

Malitha Ratnaweera ◽

Andrzej Harris ◽

Ben F. Luisi ◽

Véronique Yvette Ntsogo Enguéné

Keyword(s):

Membrane Protein ◽

Conformational Changes ◽

Drug Transport ◽

Considerable Increase ◽

Efflux Pump ◽

Transition Process ◽

Membrane Fusion Protein ◽

Allosteric Communication ◽

Inner Membrane Protein ◽

Dynamics Simulations

RND family efflux pumps are complex macromolecular machines involved in multidrug resistance by extruding antibiotics from the cell. While structural studies and molecular dynamics simulations have provided insights into the architecture and conformational states of the pumps, the path followed by conformational changes from the inner membrane protein (IMP) to the periplasmic membrane fusion protein (MFP) and to the outer membrane protein (OMP) in tripartite efflux assemblies is not fully understood. Here, we investigated AcrAB-TolC efflux pump’s allostery by comparing resting and transport states using difference distance matrices supplemented with evolutionary couplings data and buried surface area measurements. Our analysis indicated that substrate binding by the IMP triggers quaternary level conformational changes in the MFP, which induce OMP to switch from the closed state to the open state, accompanied by a considerable increase in the interface area between the MFP subunits and between the OMPs and MFPs. This suggests that the pump’s transport-ready state is at a more favourable energy level than the resting state, but raises the puzzle of how the pump does not become stably trapped in a transport-intermediate state. We propose a model for pump allostery that includes a downhill energetic transition process from a proposed ‘activated’ transport state back to the resting pump.

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Molecular Dynamics Simulations of E. coli MsbA Transmembrane Domain: Formation of a Semipore Structure

Biophysical Journal ◽

10.1529/biophysj.106.084020 ◽

2006 ◽

Vol 91 (7) ◽

pp. 2517-2531 ◽

Cited By ~ 19

Author(s):

David Y. Haubertin ◽

Hocine Madaoui ◽

Alain Sanson ◽

Raphaël Guérois ◽

Stéphane Orlowski

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Transmembrane Domain ◽

Domain Formation ◽

E Coli ◽

Dynamics Simulations

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Bonded Force Constant Derivation of Lysine-Arginine Cross-linked Advanced Glycation End-Products

10.26434/chemrxiv.5891602.v2 ◽

2018 ◽

Author(s):

Anthony Nash ◽

Nora H de Leeuw ◽

Helen L Birch

Keyword(s):

Molecular Dynamics ◽

Force Constant ◽

Computational Study ◽

Force Constants ◽

Quantum Mechanical ◽

Advanced Glycation ◽

Partial Charges ◽

Cross Links ◽

Complete Set ◽

Dynamics Simulations

<div> <div> <div> <p>The computational study of advanced glycation end-product cross- links remains largely unexplored given the limited availability of bonded force constants and equilibrium values for molecular dynamics force fields. In this article, we present the bonded force constants, atomic partial charges and equilibrium values of the arginine-lysine cross-links DOGDIC, GODIC and MODIC. The Hessian was derived from a series of <i>ab initio</i> quantum mechanical electronic structure calculations and from which a complete set of force constant and equilibrium values were generated using our publicly available software, ForceGen. Short <i>in vacuo</i> molecular dynamics simulations were performed to validate their implementation against quantum mechanical frequency calculations. </p> </div> </div> </div>

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Detection of Chemical Exchange in Methyl Groups of Macromolecules

10.26434/chemrxiv.7762067 ◽

2019 ◽

Author(s):

Michelle Gill ◽

Andrew Hsu ◽

Arthur G. Palmer, III

Keyword(s):

Relaxation Rate ◽

Chemical Exchange ◽

Double Quantum ◽

Methyl Groups ◽

Multiple Quantum ◽

Relaxation Data ◽

E Coli ◽

Methyl Trosy ◽

Dynamics Simulations ◽

Exchange Broadening

<div> <div> <div> <p>The zero- and double-quantum methyl TROSY Hahn-echo and the methyl <sup>1</sup>H-<sup>1</sup>H dipole- dipole cross-correlation nuclear magnetic resonance experiments enable estimation of multiple quantum chemical exchange broadening in methyl groups in proteins. The two relaxation rate constants are established to be linearly dependent using molecular dynamics simulations and empirical analysis of experimental data. This relationship allows chemical exchange broadening to be recognized as an increase in the Hahn-echo relaxation rate constant. The approach is illustrated by analyzing relaxation data collected at three temperatures for <i>E. coli </i>ribonuclease HI and by analyzing relaxation data collected for different cofactor and substrate complexes of <i>E. coli </i>AlkB. </p> </div> </div> </div>

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Primary and Secondary Dimer Interfaces of the Fibroblast Growth Factor Receptor 3 Transmembrane Domain: Characterization via Multiscale Molecular Dynamics Simulations

Biochemistry ◽

10.1021/bi401576k ◽

2014 ◽

Vol 53 (2) ◽

pp. 323-332 ◽

Cited By ~ 18

Author(s):

Tyler Reddy ◽

Santiago Manrique ◽

Amanda Buyan ◽

Benjamin A. Hall ◽

Alan Chetwynd ◽

...

Keyword(s):

Molecular Dynamics ◽

Growth Factor ◽

Molecular Dynamics Simulations ◽

Fibroblast Growth Factor ◽

Fibroblast Growth Factor Receptor ◽

Transmembrane Domain ◽

Growth Factor Receptor ◽

Fibroblast Growth ◽

Dynamics Simulations

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Liquid dibromomethane under pressure: a computational study

Physical Chemistry Chemical Physics ◽

10.1039/d0cp06458k ◽

2021 ◽

Vol 23 (4) ◽

pp. 2964-2971

Author(s):

Bernadeta Jasiok ◽

Mirosław Chorążewski ◽

Eugene B. Postnikov ◽

Claude Millot

Keyword(s):

Molecular Dynamics ◽

Molecular Dynamics Simulations ◽

Thermophysical Properties ◽

Computational Study ◽

Thermal Expansivity ◽

Dynamics Simulations ◽

Isobaric Thermal Expansivity

Thermophysical properties of liquid dibromomethane are investigated by molecular dynamics simulations between 268 and 328 K at pressures up to 3000 bar. Notably, the isotherms of the isobaric thermal expansivity cross around 800 bar.

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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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Bioactive constituents with antibacterial, resistance modulation, anti-biofilm formation and efflux pump inhibition properties from Aidia genipiflora stem bark

Clinical Phytoscience ◽

10.1186/s40816-021-00266-4 ◽

2021 ◽

Vol 7 (1) ◽

Author(s):

Daniel Anokwah ◽

Evelyn Asante-Kwatia ◽

Abraham Y. Mensah ◽

Cynthia Amaning Danquah ◽

Benjamin K. Harley ◽

...

Keyword(s):

Biofilm Formation ◽

Efflux Pump ◽

Ethyl Acetate Fraction ◽

Stem Bark ◽

Resistance Mechanisms ◽

Bioactive Constituents ◽

Biofilm Inhibition ◽

E Coli ◽

Efflux Pump Inhibition ◽

Oleanonic Acid

Abstract Background Antimicrobial resistance is a global health challenge. The involvement of bacterial biofilms and efflux pumps in the development of multidrug resistance (MDR) is well established. Medicinal plants have been proposed as alternatives for combating MDR focusing on their bioactive constituents with resistance modulatory activities. This study was aimed at investigating the stem bark of Aidia genipiflora for bioactive constituents with anti-biofilm, efflux pump inhibition and resistance modulatory activities. Method The crude methanol extract was purified by column chromatography and isolated compounds characterized by mass and nuclear magnetic resonance spectrometry. Antibacterial activity was determined by the High-throughput spot culture growth inhibition and the broth micro-dilution assay. The ethidium bromide accumulation assay was used to determine efflux pump inhibition property. Biofilm inhibition was determined in a microplate crystal violet retention assay. Results Purification of the ethyl acetate fraction led to the isolation of oleanonic acid (1), 4-hydroxy cinnamic acid docosyl ester (2), β-stigmasterol/β-sitosterol (mixture 3a/b) and D-mannitol (4). The minimum inhibitory concentrations (MICs) ranged from 250 to > 500 μg/mL for extracts and fractions and from 15 to 250 μg/mL for compounds. In the presence of sub-inhibitory concentrations of the compounds, the MIC of amoxicillin against E. coli (20 μg/mL) and P. aeruginosa (320 μg/mL) was reduced by 32 and 10 folds respectively. The whole extract demonstrated anti-biofilm formation and efflux pump inhibition in E. coli, S. aureus and P. aeruginosa. The sterol mixture (3a/b) at concentration of 100 μg/mL caused the highest inhibition (73%) of biofilm formation in S. aureus. Oleanonic acid (1) demonstrated remarkable efflux pump inhibition at MIC of 7.8 μg/mL in E. coli better than the standard drugs verapamil and chlorpromazine. Conclusion This study confirms the prospects of A. genipiflora as a source of new antibacterial agents and adjuvants that could interact with some resistance mechanisms in bacteria to enhance the activity of hitherto ineffective antibiotics. “A small portion of the study has been presented in a conference in the form of poster”.

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