Identification of Binding Sites for Efflux Pump Inhibitors of the AcrAB-TolC Component AcrA

Zbigniew M. Darzynkiewicz; Adam T. Green; Narges Abdali; Anthony Hazel; Ronnie L. Fulton; Joseph Kimball; Zygmunt Gryczynski; James C. Gumbart; Jerry M. Parks; Jeremy C. Smith; Helen I. Zgurskaya

doi:10.1016/j.bpj.2019.01.010

Identification of Binding Sites for Efflux Pump Inhibitors of the AcrAB-TolC Component AcrA

Zbigniew M. Darzynkiewicz, Adam T. Green, Narges Abdali, Anthony Hazel, Ronnie L. Fulton, Joseph Kimball, Zygmunt Gryczynski, James C. Gumbart, Jerry M. Parks, Jeremy C. Smith, Helen I. Zgurskaya

Molecular Biophysics

Research output: Contribution to journal › Article › peer-review

23 Scopus citations

Abstract

The overexpression of multidrug efflux pumps is an important mechanism of clinical resistance in Gram-negative bacteria. Recently, four small molecules were discovered that inhibit efflux in Escherichia coli and interact with the AcrAB-TolC efflux pump component AcrA. However, the binding site(s) for these molecules was not determined. Here, we combine ensemble docking and molecular dynamics simulations with tryptophan fluorescence spectroscopy, site-directed mutagenesis, and antibiotic susceptibility assays to probe binding sites and effects of binding of these molecules. We conclude that clorobiocin and SLU-258 likely bind at a site located between the lipoyl and β-barrel domains of AcrA.

Original language	English
Pages (from-to)	648-658
Number of pages	11
Journal	Biophysical Journal
Volume	116
Issue number	4
DOIs	https://doi.org/10.1016/j.bpj.2019.01.010
State	Published - Feb 19 2019

Funding

This work was supported by National Institutes of Health, United States grant AI052293 to H.I.Z. This work used resources of the Compute and Data Environment for Science at Oak Ridge National Laboratory, which is managed by UT-Battelle for the Department of Energy, United States under contract no. DE-AC05-00OR22725 . A.H. and J.C.G. acknowledge support from the National Science Foundation, United States through grant MCB-1452464 as well as the Extreme Science and Engineering Discovery Environment , which is supported by National Science Foundation grant number OCI-1053575 . This work was supported by National Institutes of Health, United States grant AI052293 to H.I.Z. This work used resources of the Compute and Data Environment for Science at Oak Ridge National Laboratory, which is managed by UT-Battelle for the Department of Energy, United States under contract no. DE-AC05-00OR22725. A.H. and J.C.G. acknowledge support from the National Science Foundation, United States through grant MCB-1452464 as well as the Extreme Science and Engineering Discovery Environment, which is supported by National Science Foundation grant number OCI-1053575.

Funders	Funder number
UT-Battelle
National Science Foundation	MCB-1452464, OCI-1053575
National Institutes of Health	AI052293
U.S. Department of Energy	DE-AC05-00OR22725

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10.1016/j.bpj.2019.01.010

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title = "Identification of Binding Sites for Efflux Pump Inhibitors of the AcrAB-TolC Component AcrA",

abstract = "The overexpression of multidrug efflux pumps is an important mechanism of clinical resistance in Gram-negative bacteria. Recently, four small molecules were discovered that inhibit efflux in Escherichia coli and interact with the AcrAB-TolC efflux pump component AcrA. However, the binding site(s) for these molecules was not determined. Here, we combine ensemble docking and molecular dynamics simulations with tryptophan fluorescence spectroscopy, site-directed mutagenesis, and antibiotic susceptibility assays to probe binding sites and effects of binding of these molecules. We conclude that clorobiocin and SLU-258 likely bind at a site located between the lipoyl and β-barrel domains of AcrA.",

author = "Darzynkiewicz, {Zbigniew M.} and Green, {Adam T.} and Narges Abdali and Anthony Hazel and Fulton, {Ronnie L.} and Joseph Kimball and Zygmunt Gryczynski and Gumbart, {James C.} and Parks, {Jerry M.} and Smith, {Jeremy C.} and Zgurskaya, {Helen I.}",

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doi = "10.1016/j.bpj.2019.01.010",

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T1 - Identification of Binding Sites for Efflux Pump Inhibitors of the AcrAB-TolC Component AcrA

AU - Darzynkiewicz, Zbigniew M.

AU - Green, Adam T.

AU - Abdali, Narges

AU - Hazel, Anthony

AU - Fulton, Ronnie L.

AU - Kimball, Joseph

AU - Gryczynski, Zygmunt

AU - Gumbart, James C.

AU - Parks, Jerry M.

AU - Smith, Jeremy C.

AU - Zgurskaya, Helen I.

PY - 2019/2/19

Y1 - 2019/2/19

N2 - The overexpression of multidrug efflux pumps is an important mechanism of clinical resistance in Gram-negative bacteria. Recently, four small molecules were discovered that inhibit efflux in Escherichia coli and interact with the AcrAB-TolC efflux pump component AcrA. However, the binding site(s) for these molecules was not determined. Here, we combine ensemble docking and molecular dynamics simulations with tryptophan fluorescence spectroscopy, site-directed mutagenesis, and antibiotic susceptibility assays to probe binding sites and effects of binding of these molecules. We conclude that clorobiocin and SLU-258 likely bind at a site located between the lipoyl and β-barrel domains of AcrA.

AB - The overexpression of multidrug efflux pumps is an important mechanism of clinical resistance in Gram-negative bacteria. Recently, four small molecules were discovered that inhibit efflux in Escherichia coli and interact with the AcrAB-TolC efflux pump component AcrA. However, the binding site(s) for these molecules was not determined. Here, we combine ensemble docking and molecular dynamics simulations with tryptophan fluorescence spectroscopy, site-directed mutagenesis, and antibiotic susceptibility assays to probe binding sites and effects of binding of these molecules. We conclude that clorobiocin and SLU-258 likely bind at a site located between the lipoyl and β-barrel domains of AcrA.

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JO - Biophysical Journal

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Identification of Binding Sites for Efflux Pump Inhibitors of the AcrAB-TolC Component AcrA

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