Lpp positions peptidoglycan at the AcrA-TolC interface in the AcrAB-TolC multidrug efflux pump

James C. Gumbart; Josie L. Ferreira; Hyea Hwang; Anthony J. Hazel; Connor J. Cooper; Jerry M. Parks; Jeremy C. Smith; Helen I. Zgurskaya; Morgan Beeby

doi:10.1016/j.bpj.2021.08.016

Lpp positions peptidoglycan at the AcrA-TolC interface in the AcrAB-TolC multidrug efflux pump

James C. Gumbart, Josie L. Ferreira, Hyea Hwang, Anthony J. Hazel, Connor J. Cooper, Jerry M. Parks, Jeremy C. Smith, Helen I. Zgurskaya, Morgan Beeby

Molecular Biophysics

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

14 Scopus citations

Abstract

The multidrug efflux pumps of Gram-negative bacteria are a class of complexes that span the periplasm, coupling both the inner and outer membranes to expel toxic molecules. The best-characterized example of these tripartite pumps is the AcrAB-TolC complex of Escherichia coli. However, how the complex interacts with the peptidoglycan (PG) cell wall, which is anchored to the outer membrane (OM) by Braun's lipoprotein (Lpp), is still largely unknown. In this work, we present molecular dynamics simulations of a complete, atomistic model of the AcrAB-TolC complex with the inner membrane, OM, and PG layers all present. We find that the PG localizes to the junction of AcrA and TolC, in agreement with recent cryo-tomography data. Free-energy calculations reveal that the positioning of PG is determined by the length and conformation of multiple Lpp copies anchoring it to the OM. The distance between the PG and OM measured in cryo-electron microscopy images of wild-type E. coli also agrees with the simulation-derived spacing. Sequence analysis of AcrA suggests a conserved role for interactions with PG in the assembly and stabilization of efflux pumps, one that may extend to other trans-envelope complexes as well.

Original language	English
Pages (from-to)	3973-3982
Number of pages	10
Journal	Biophysical Journal
Volume	120
Issue number	18
DOIs	https://doi.org/10.1016/j.bpj.2021.08.016
State	Published - Sep 21 2021

Funding

This work was supported by National Institutes of Health grant R01-AI052293 to H.I.Z. J.M.P. J.C.S. and J.C.G. This work was also supported by a Medical Research Council grant MR/P019374/1 to M.B. and a Medical Research Council PhD Doctoral Training Partnership award grant number MR/K501281/1 to J.L.F. An award of computer time was provided by the Innovative and Novel Computational Impact on Theory and Experiment (INCITE) program (BIP150). This research used resources of the Oak Ridge Leadership Computing Facility at Oak Ridge National Laboratory and the Compute and Data Environment for Science (CADES), which is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC05-00OR22725. Additional computational resources were provided through the Extreme Science and Engineering Discovery Environment (TG-MCB130173), which is supported by National Science Foundation grant ACI-1548562. This work was supported by National Institutes of Health grant R01-AI052293 to H.I.Z., J.M.P., J.C.S., and J.C.G. This work was also supported by a Medical Research Council grant MR/P019374/1 to M.B. and a Medical Research Council PhD Doctoral Training Partnership award grant number MR/K501281/1 to J.L.F. An award of computer time was provided by the Innovative and Novel Computational Impact on Theory and Experiment (INCITE) program (BIP150). This research used resources of the Oak Ridge Leadership Computing Facility at Oak Ridge National Laboratory and the Compute and Data Environment for Science (CADES), which is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC05-00OR22725 . Additional computational resources were provided through the Extreme Science and Engineering Discovery Environment ( TG-MCB130173 ), which is supported by National Science Foundation grant ACI-1548562 .

Funders	Funder number
Compute and Data Environment for Science
Extreme Science and Engineering Discovery Environment	TG-MCB130173
National Science Foundation	ACI-1548562
National Institutes of Health	R01-AI052293
U.S. Department of Energy	DE-AC05-00OR22725
Office of Science
Oak Ridge National Laboratory
Medical Research Council	MR/K501281/1, MR/P019374/1, BIP150

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title = "Lpp positions peptidoglycan at the AcrA-TolC interface in the AcrAB-TolC multidrug efflux pump",

abstract = "The multidrug efflux pumps of Gram-negative bacteria are a class of complexes that span the periplasm, coupling both the inner and outer membranes to expel toxic molecules. The best-characterized example of these tripartite pumps is the AcrAB-TolC complex of Escherichia coli. However, how the complex interacts with the peptidoglycan (PG) cell wall, which is anchored to the outer membrane (OM) by Braun's lipoprotein (Lpp), is still largely unknown. In this work, we present molecular dynamics simulations of a complete, atomistic model of the AcrAB-TolC complex with the inner membrane, OM, and PG layers all present. We find that the PG localizes to the junction of AcrA and TolC, in agreement with recent cryo-tomography data. Free-energy calculations reveal that the positioning of PG is determined by the length and conformation of multiple Lpp copies anchoring it to the OM. The distance between the PG and OM measured in cryo-electron microscopy images of wild-type E. coli also agrees with the simulation-derived spacing. Sequence analysis of AcrA suggests a conserved role for interactions with PG in the assembly and stabilization of efflux pumps, one that may extend to other trans-envelope complexes as well.",

author = "Gumbart, {James C.} and Ferreira, {Josie L.} and Hyea Hwang and Hazel, {Anthony J.} and Cooper, {Connor J.} and Parks, {Jerry M.} and Smith, {Jeremy C.} and Zgurskaya, {Helen I.} and Morgan Beeby",

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AU - Gumbart, James C.

AU - Ferreira, Josie L.

AU - Hwang, Hyea

AU - Hazel, Anthony J.

AU - Cooper, Connor J.

AU - Parks, Jerry M.

AU - Smith, Jeremy C.

AU - Zgurskaya, Helen I.

AU - Beeby, Morgan

PY - 2021/9/21

Y1 - 2021/9/21

N2 - The multidrug efflux pumps of Gram-negative bacteria are a class of complexes that span the periplasm, coupling both the inner and outer membranes to expel toxic molecules. The best-characterized example of these tripartite pumps is the AcrAB-TolC complex of Escherichia coli. However, how the complex interacts with the peptidoglycan (PG) cell wall, which is anchored to the outer membrane (OM) by Braun's lipoprotein (Lpp), is still largely unknown. In this work, we present molecular dynamics simulations of a complete, atomistic model of the AcrAB-TolC complex with the inner membrane, OM, and PG layers all present. We find that the PG localizes to the junction of AcrA and TolC, in agreement with recent cryo-tomography data. Free-energy calculations reveal that the positioning of PG is determined by the length and conformation of multiple Lpp copies anchoring it to the OM. The distance between the PG and OM measured in cryo-electron microscopy images of wild-type E. coli also agrees with the simulation-derived spacing. Sequence analysis of AcrA suggests a conserved role for interactions with PG in the assembly and stabilization of efflux pumps, one that may extend to other trans-envelope complexes as well.

AB - The multidrug efflux pumps of Gram-negative bacteria are a class of complexes that span the periplasm, coupling both the inner and outer membranes to expel toxic molecules. The best-characterized example of these tripartite pumps is the AcrAB-TolC complex of Escherichia coli. However, how the complex interacts with the peptidoglycan (PG) cell wall, which is anchored to the outer membrane (OM) by Braun's lipoprotein (Lpp), is still largely unknown. In this work, we present molecular dynamics simulations of a complete, atomistic model of the AcrAB-TolC complex with the inner membrane, OM, and PG layers all present. We find that the PG localizes to the junction of AcrA and TolC, in agreement with recent cryo-tomography data. Free-energy calculations reveal that the positioning of PG is determined by the length and conformation of multiple Lpp copies anchoring it to the OM. The distance between the PG and OM measured in cryo-electron microscopy images of wild-type E. coli also agrees with the simulation-derived spacing. Sequence analysis of AcrA suggests a conserved role for interactions with PG in the assembly and stabilization of efflux pumps, one that may extend to other trans-envelope complexes as well.

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Lpp positions peptidoglycan at the AcrA-TolC interface in the AcrAB-TolC multidrug efflux pump

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