An allosteric transport mechanism for the AcrAB-TolC Multidrug Efflux Pump

  1. Zhao Wang
  2. Guizhen Fan
  3. Corey F Hryc
  4. James N Blaza
  5. Irina I Serysheva
  6. Michael F Schmid
  7. Wah Chiu  Is a corresponding author
  8. Ben F Luisi  Is a corresponding author
  9. Dijun Du  Is a corresponding author
  1. Baylor College of Medicine, United States
  2. The University of Texas Health Science Center at Houston Medical School, United States
  3. MRC Mitochondrial Biology Unit, United Kingdom
  4. University of Cambridge, United Kingdom

Abstract

Bacterial efflux pumps confer multidrug resistance by transporting diverse antibiotics from the cell. In Gram-negative bacteria, some of these pumps form multi-protein assemblies that span the cell envelope. Here we report the near-atomic resolution cryoEM structures of the Escherichia coli AcrAB-TolC multidrug efflux pump in resting and drug transport states, revealing a quaternary structural switch that allosterically couples and synchronizes initial ligand binding with channel opening. Within the transport-activated state, the channel remains open even though the pump cycles through three distinct conformations. Collectively, our data provide a dynamic mechanism for the assembly and operation of the AcrAB-TolC pump.

Data availability

The following data sets were generated
    1. D. Du
    2. B.F. Luisi
    (2017) Crystal structure of AcrBZ complex: 2017
    Publicly available at PDB (accession no: 5NC5).

Article and author information

Author details

  1. Zhao Wang

    National Center for Macromolecular Imaging, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Guizhen Fan

    Department of Biochemistry and Molecular Biology, The University of Texas Health Science Center at Houston Medical School, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Corey F Hryc

    National Center for Macromolecular Imaging, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. James N Blaza

    MRC Mitochondrial Biology Unit, Cambridge, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5420-2116
  5. Irina I Serysheva

    Department of Biochemistry and Molecular Biology, The University of Texas Health Science Center at Houston Medical School, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Michael F Schmid

    National Center for Macromolecular Imaging, Baylor College of Medicine, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Wah Chiu

    National Center for Macromolecular Imaging, Baylor College of Medicine, Houston, United States
    For correspondence
    wah@bcm.edu
    Competing interests
    The authors declare that no competing interests exist.
  8. Ben F Luisi

    Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom
    For correspondence
    bfl20@cam.ac.uk
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1144-9877
  9. Dijun Du

    Department of Biochemistry, University of Cambridge, Cambridge, United Kingdom
    For correspondence
    dd339@cam.ac.uk
    Competing interests
    The authors declare that no competing interests exist.

Funding

Wellcome

  • Ben F Luisi

Human Frontier Science Program

  • Ben F Luisi

National Institutes of Health (P41GM103832)

  • Wah Chiu

American Heart Association (16GRNT29720001)

  • Irina I Serysheva

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2017, Wang et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 13,427
    views
  • 1,969
    downloads
  • 202
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Zhao Wang
  2. Guizhen Fan
  3. Corey F Hryc
  4. James N Blaza
  5. Irina I Serysheva
  6. Michael F Schmid
  7. Wah Chiu
  8. Ben F Luisi
  9. Dijun Du
(2017)
An allosteric transport mechanism for the AcrAB-TolC Multidrug Efflux Pump
eLife 6:e24905.
https://doi.org/10.7554/eLife.24905

Share this article

https://doi.org/10.7554/eLife.24905

Further reading

    1. Immunology and Inflammation
    2. Structural Biology and Molecular Biophysics
    Douwe Schulte, Marta Šiborová ... Joost Snijder
    Research Article

    Antibodies are a major component of adaptive immunity against invading pathogens. Here, we explore possibilities for an analytical approach to characterize the antigen-specific antibody repertoire directly from the secreted proteins in convalescent serum. This approach aims to perform simultaneous antibody sequencing and epitope mapping using a combination of single particle cryo-electron microscopy (cryoEM) and bottom-up proteomics techniques based on mass spectrometry (LC-MS/MS). We evaluate the performance of the deep-learning tool ModelAngelo in determining de novo antibody sequences directly from reconstructed 3D volumes of antibody-antigen complexes. We demonstrate that while map quality is a critical bottleneck, it is possible to sequence antibody variable domains from cryoEM reconstructions with accuracies of up to 80–90%. While the rate of errors exceeds the typical levels of somatic hypermutation, we show that the ModelAngelo-derived sequences can be used to assign the used V-genes. This provides a functional guide to assemble de novo peptides from LC-MS/MS data more accurately and improves the tolerance to a background of polyclonal antibody sequences. Following this proof-of-principle, we discuss the feasibility and future directions of this approach to characterize antigen-specific antibody repertoires.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Yamato Niitani, Kohei Matsuzaki ... Michio Tomishige
    Research Article

    The two identical motor domains (heads) of dimeric kinesin-1 move in a hand-over-hand process along a microtubule, coordinating their ATPase cycles such that each ATP hydrolysis is tightly coupled to a step and enabling the motor to take many steps without dissociating. The neck linker, a structural element that connects the two heads, has been shown to be essential for head–head coordination; however, which kinetic step(s) in the chemomechanical cycle is ‘gated’ by the neck linker remains unresolved. Here, we employed pre-steady-state kinetics and single-molecule assays to investigate how the neck-linker conformation affects kinesin’s motility cycle. We show that the backward-pointing configuration of the neck linker in the front kinesin head confers higher affinity for microtubule, but does not change ATP binding and dissociation rates. In contrast, the forward-pointing configuration of the neck linker in the rear kinesin head decreases the ATP dissociation rate but has little effect on microtubule dissociation. In combination, these conformation-specific effects of the neck linker favor ATP hydrolysis and dissociation of the rear head prior to microtubule detachment of the front head, thereby providing a kinetic explanation for the coordinated walking mechanism of dimeric kinesin.