1. Microbiology and Infectious Disease
  2. Structural Biology and Molecular Biophysics

Membrane insertion of α-xenorhabdolysin in near-atomic detail

  1. Evelyn Schubert
  2. Ingrid R Vetter
  3. Daniel Prumbaum
  4. Pawel A Penczek
  5. Stefan Raunser  Is a corresponding author
  1. Max Planck Institute of Molecular Physiology, Germany
  2. The University of Texas, United States
https://doi.org/10.7554/eLife.38017
Share this article
Cite this article
  1. Evelyn Schubert
  2. Ingrid R Vetter
  3. Daniel Prumbaum
  4. Pawel A Penczek
  5. Stefan Raunser
(2018)
Membrane insertion of α-xenorhabdolysin in near-atomic detail
eLife 7:e38017.
https://doi.org/10.7554/eLife.38017
  2. Download BibTeX
  3. Download .RIS

Abstract

α-Xenorhabdolysins (Xax) are α-pore-forming toxins (α-PFT) that form 1-1.3 MDa large pore complexes to perforate the host cell membrane. PFTs are used by a variety of bacterial pathogens to attack host cells. Due to the lack of structural information, the molecular mechanism of action of Xax toxins is poorly understood. Here, we report the cryo-EM structure of the XaxAB pore complex from Xenorhabdus nematophila and the crystal structures of the soluble monomers of XaxA and XaxB. The structures reveal that XaxA and XaxB are built similarly and appear as heterodimers in the 12-15 subunits containing pore, classifying XaxAB as bi-component α-PFT. Major conformational changes in XaxB, including the swinging out of an amphipathic helix are responsible for membrane insertion. XaxA acts as an activator and stabilizer for XaxB that forms the actual transmembrane pore. Based on our results, we propose a novel structural model for the mechanism of Xax intoxication.

Data availability

The electron density map after post-processing has been deposited to the EMDB under accession code EMD-0088. The final model of XaxAB was submitted to the PDB under the accession code 6GY6. Coordinates of XaxA and XaxB in the soluble form have been deposited in the Protein Data Bank under accession codes PDB 6GY8 and PDB 6G47.

The following data sets were generated
    1. Raunser S
    2. Schubert E
    (2018) Crystal structure of XaxA from Xenorhabdus nematophila
    Publicly available at the RCSB Protein Data Bank (accession no: 6GY8).
    https://www.rcsb.org
    1. Raunser S
    2. Schubert E
    (2018) XaxAB pore complex from Xenorhabdus nematophila
    Publicly available at the RCSB Protein Data Bank (accession no: EMD-0088, PDB ID 6GY6).
    https://www.rcsb.org
    1. Raunser S
    2. Schubert E
    (2018) Crystal structure of XaxB from Xenorhabdus nematophila
    Publicly available at the RCSB Protein Data Bank (accession no: 6GY7.
    https://www.rcsb.org

Article and author information

Author details

  1. Evelyn Schubert

    Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany
    Competing interests
    The authors declare that no competing interests exist.

  2. Ingrid R Vetter

    Department of Mechanistic Cell Biology, Max Planck Institute of Molecular Physiology, Dortmund, Germany
    Competing interests
    The authors declare that no competing interests exist.

  3. Daniel Prumbaum

    Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany
    Competing interests
    The authors declare that no competing interests exist.

  4. Pawel A Penczek

    Department of Biochemistry and Molecular Biology, The University of Texas, Houston, United States
    Competing interests
    The authors declare that no competing interests exist.

  5. Stefan Raunser

    Department of Structural Biochemistry, Max Planck Institute of Molecular Physiology, Dortmund, Germany
    For correspondence
    stefan.raunser@mpi-dortmund.mpg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9373-3016

Funding

Max-Planck-Gesellschaft (Open-access funding)

  • Stefan Raunser

European Commission

  • Stefan Raunser

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 Medicine, United States

Version history

  1. Received: May 2, 2018
  2. Accepted: July 15, 2018
  3. Accepted Manuscript published: July 16, 2018 (version 1)
  4. Version of Record published: August 10, 2018 (version 2)

Copyright

© 2018, Schubert 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

  • 2,618
    views
  • 441
    downloads
  • 25
    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. Evelyn Schubert
  2. Ingrid R Vetter
  3. Daniel Prumbaum
  4. Pawel A Penczek
  5. Stefan Raunser
(2018)
Membrane insertion of α-xenorhabdolysin in near-atomic detail
eLife 7:e38017.
https://doi.org/10.7554/eLife.38017

Share this article

https://doi.org/10.7554/eLife.38017

Categories and tags

Further reading

    1. Microbiology and Infectious Disease

    The target of rapamycin signaling pathway regulates vegetative development, aflatoxin biosynthesis, and pathogenicity in Aspergillus flavus

    Guoqi Li, Xiaohong Cao ... Shihua Wang
    Research Article

    The target of rapamycin (TOR) signaling pathway is highly conserved and plays a crucial role in diverse biological processes in eukaryotes. Despite its significance, the underlying mechanism of the TOR pathway in Aspergillus flavus remains elusive. In this study, we comprehensively analyzed the TOR signaling pathway in A. flavus by identifying and characterizing nine genes that encode distinct components of this pathway. The FK506-binding protein Fkbp3 and its lysine succinylation are important for aflatoxin production and rapamycin resistance. The TorA kinase plays a pivotal role in the regulation of growth, spore production, aflatoxin biosynthesis, and responses to rapamycin and cell membrane stress. As a significant downstream effector molecule of the TorA kinase, the Sch9 kinase regulates aflatoxin B1 (AFB1) synthesis, osmotic and calcium stress response in A. flavus, and this regulation is mediated through its S_TKc, S_TK_X domains, and the ATP-binding site at K340. We also showed that the Sch9 kinase may have a regulatory impact on the high osmolarity glycerol (HOG) signaling pathway. TapA and TipA, the other downstream components of the TorA kinase, play a significant role in regulating cell wall stress response in A. flavus. Moreover, the members of the TapA-phosphatase complexes, SitA and Ppg1, are important for various biological processes in A. flavus, including vegetative growth, sclerotia formation, AFB1 biosynthesis, and pathogenicity. We also demonstrated that SitA and Ppg1 are involved in regulating lipid droplets (LDs) biogenesis and cell wall integrity (CWI) signaling pathways. In addition, another phosphatase complex, Nem1/Spo7, plays critical roles in hyphal development, conidiation, aflatoxin production, and LDs biogenesis. Collectively, our study has provided important insight into the regulatory network of the TOR signaling pathway and has elucidated the underlying molecular mechanisms of aflatoxin biosynthesis in A. flavus.

    1. Microbiology and Infectious Disease

    Bacteria are a major determinant of Orsay virus transmission and infection in Caenorhabditis elegans

    Brian G Vassallo, Noemie Scheidel ... Dennis H Kim
    Research Article

    The microbiota is a key determinant of the physiology and immunity of animal hosts. The factors governing the transmissibility of viruses between susceptible hosts are incompletely understood. Bacteria serve as food for Caenorhabditis elegans and represent an integral part of the natural environment of C. elegans. We determined the effects of bacteria isolated with C. elegans from its natural environment on the transmission of Orsay virus in C. elegans using quantitative virus transmission and host susceptibility assays. We observed that Ochrobactrum species promoted Orsay virus transmission, whereas Pseudomonas lurida MYb11 attenuated virus transmission relative to the standard laboratory bacterial food Escherichia coli OP50. We found that pathogenic Pseudomonas aeruginosa strains PA01 and PA14 further attenuated virus transmission. We determined that the amount of Orsay virus required to infect 50% of a C. elegans population on P. lurida MYb11 compared with Ochrobactrum vermis MYb71 was dramatically increased, over three orders of magnitude. Host susceptibility was attenuated even further in the presence of P. aeruginosa PA14. Genetic analysis of the determinants of P. aeruginosa required for attenuation of C. elegans susceptibility to Orsay virus infection revealed a role for regulators of quorum sensing. Our data suggest that distinct constituents of the C. elegans microbiota and potential pathogens can have widely divergent effects on Orsay virus transmission, such that associated bacteria can effectively determine host susceptibility versus resistance to viral infection. Our study provides quantitative evidence for a critical role for tripartite host-virus-bacteria interactions in determining the transmissibility of viruses among susceptible hosts.

    1. Biochemistry and Chemical Biology
    2. Microbiology and Infectious Disease

    Chemoproteomics validates selective targeting of Plasmodium M1 alanyl aminopeptidase as an antimalarial strategy

    Carlo Giannangelo, Matthew P Challis ... Darren J Creek
    Research Article

    New antimalarial drug candidates that act via novel mechanisms are urgently needed to combat malaria drug resistance. Here, we describe the multi-omic chemical validation of Plasmodium M1 alanyl metalloaminopeptidase as an attractive drug target using the selective inhibitor, MIPS2673. MIPS2673 demonstrated potent inhibition of recombinant Plasmodium falciparum (PfA-M1) and Plasmodium vivax (PvA-M1) M1 metalloaminopeptidases, with selectivity over other Plasmodium and human aminopeptidases, and displayed excellent in vitro antimalarial activity with no significant host cytotoxicity. Orthogonal label-free chemoproteomic methods based on thermal stability and limited proteolysis of whole parasite lysates revealed that MIPS2673 solely targets PfA-M1 in parasites, with limited proteolysis also enabling estimation of the binding site on PfA-M1 to within ~5 Å of that determined by X-ray crystallography. Finally, functional investigation by untargeted metabolomics demonstrated that MIPS2673 inhibits the key role of PfA-M1 in haemoglobin digestion. Combined, our unbiased multi-omic target deconvolution methods confirmed the on-target activity of MIPS2673, and validated selective inhibition of M1 alanyl metalloaminopeptidase as a promising antimalarial strategy.