1. Structural Biology and Molecular Biophysics
Download icon

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
Research Article
  • Cited 114
  • Views 8,259
  • Annotations
Cite this article as: eLife 2017;6:e24905 doi: 10.7554/eLife.24905

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).
    http://www.wwpdb.org

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.

Reviewing Editor

  1. Olga Boudker, Weill Cornell Medical College, United States

Publication history

  1. Received: January 5, 2017
  2. Accepted: March 14, 2017
  3. Accepted Manuscript published: March 29, 2017 (version 1)
  4. Version of Record published: April 25, 2017 (version 2)

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

  • 8,259
    Page views
  • 1,479
    Downloads
  • 114
    Citations

Article citation count generated by polling the highest count across the following sources: Scopus, Crossref, PubMed Central.

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)

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

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

Categories and tags

Research organism

Further reading

    1. Immunology and Inflammation
    2. Structural Biology and Molecular Biophysics

    Neutrophil-mediated oxidative stress and albumin structural damage predict COVID-19-associated mortality

    Mohamed A Badawy et al.
    Research Article

    Human serum albumin (HSA) is the frontline antioxidant protein in blood with established anti-inflammatory and anticoagulation functions. Here we report that COVID-19-induced oxidative stress inflicts structural damages to HSA and is linked with mortality outcome in critically ill patients. We recruited 39 patients who were followed up for a median of 12.5 days (1-35 days), among them 23 had died. Analyzing blood samples from patients and healthy individuals (n=11), we provide evidence that neutrophils are major sources of oxidative stress in blood and that hydrogen peroxide is highly accumulated in plasmas of non-survivors. We then analyzed electron paramagnetic resonance (EPR) spectra of spin labelled fatty acids (SLFA) bound with HSA in whole blood of control, survivor, and non-survivor subjects (n=10-11). Non-survivor' HSA showed dramatically reduced protein packing order parameter, faster SLFA correlational rotational time, and smaller S/W ratio (strong-binding/weak-binding sites within HSA), all reflecting remarkably fluid protein microenvironments. Following loading/unloading of 16-DSA we show that transport function of HSA maybe impaired in severe patients. Stratified at the means, Kaplan–Meier survival analysis indicated that lower values of S/W ratio and accumulated H2O2 in plasma significantly predicted in-hospital mortality (S/W≤0.15, 81.8% (18/22) vs. S/W>0.15, 18.2% (4/22), p=0.023; plasma [H2O2]>8.6 mM, 65.2% (15/23) vs. 34.8% (8/23), p=0.043). When we combined these two parameters as the ratio ((S/W)/[H2O2]) to derive a risk score, the resultant risk score lower than the mean (< 0.019) predicted mortality with high fidelity (95.5% (21/22) vs. 4.5% (1/22), logrank c2 = 12.1, p=4.9x10-4). The derived parameters may provide a surrogate marker to assess new candidates for COVID-19 treatments targeting HSA replacements and/or oxidative stress.

    1. Cell Biology
    2. Structural Biology and Molecular Biophysics

    EGFR transactivates RON to drive oncogenic crosstalk

    Carolina Franco Nitta et al.
    Research Article

    Crosstalk between different receptor tyrosine kinases (RTKs) is thought to drive oncogenic signaling and allow therapeutic escape. EGFR and RON are two such RTKs from different subfamilies, which engage in crosstalk through unknown mechanisms. We combined high-resolution imaging with biochemical and mutational studies to ask how EGFR and RON communicate. EGF stimulation promotes EGFR-dependent phosphorylation of RON, but ligand stimulation of RON does not trigger EGFR phosphorylation – arguing that crosstalk is unidirectional. Nanoscale imaging reveals association of EGFR and RON in common plasma membrane microdomains. Two-color single particle tracking captured formation of complexes between RON and EGF-bound EGFR. Our results further show that RON is a substrate for EGFR kinase, and that transactivation of RON requires formation of a signaling competent EGFR dimer. These results support a role for direct EGFR/RON interactions in propagating crosstalk, such that EGF-stimulated EGFR phosphorylates RON to activate RON-directed signaling.

    1. Structural Biology and Molecular Biophysics

    Structure of Mycobacterium tuberculosis cytochrome bcc in complex with Q203 and TB47, two anti-TB drug candidates

    Shan Zhou et al.
    Research Article

    Pathogenic mycobacteria pose a sustained threat to global human health. Recently, cytochrome bcc complexes have gained interest as targets for antibiotic drug development. However, there is currently no structural information for the cytochrome bcc complex from these pathogenic mycobacteria. Here, we report the structures of Mycobacterium tuberculosis cytochrome bcc alone (2.68 Å resolution) and in complex with clinical drug candidates Q203 (2.67 Å resolution) and TB47 (2.93 Å resolution) determined by single-particle cryo-electron microscopy. M. tuberculosis cytochrome bcc forms a dimeric assembly with endogenous menaquinone/menaquinol bound at the quinone/quinol-binding pockets. We observe Q203 and TB47 bound at the quinol-binding site and stabilized by hydrogen bonds with the side chains of QcrBThr313 and QcrBGlu314, residues that are conserved across pathogenic mycobacteria. These high-resolution images provide a basis for the design of new mycobacterial cytochrome bcc inhibitors that could be developed into broad-spectrum drugs to treat mycobacterial infections.