Home LiteratureArticle Details
PMID: 25778700 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, Non-P.H.S.

Atomic structure of anthrax protective antigen pore elucidates toxin translocation.

Nature ·Vol. 521 ·No. 7553 ·2015-05-28 ·Pages 545-9

Jiang J, Pentelute BL, Collier RJ, Zhou ZH

Abstract

Anthrax toxin, comprising protective antigen, lethal factor, and oedema factor, is the major virulence factor of Bacillus anthracis, an agent that causes high mortality in humans and animals. Protective antigen forms oligomeric prepores that undergo conversion to membrane-spanning pores by endosomal acidification, and these pores translocate the enzymes lethal factor and oedema factor into the cytosol of target cells. Protective antigen is not only a vaccine component and therapeutic target for anthrax infections but also an excellent model system for understanding the mechanism of protein translocation. On the basis of biochemical and electrophysiological results, researchers have proposed that a phi (Φ)-clamp composed of phenylalanine (Phe)427 residues of protective antigen catalyses protein translocation via a charge-state-dependent Brownian ratchet. Although atomic structures of protective antigen prepores are available, how protective antigen senses low pH, converts to active pore, and translocates lethal factor and oedema factor are not well defined without an atomic model of its pore. Here, by cryo-electron microscopy with direct electron counting, we determine the protective antigen pore structure at 2.9-Å resolution. The structure reveals the long-sought-after catalytic Φ-clamp and the membrane-spanning translocation channel, and supports the Brownian ratchet model for protein translocation. Comparisons of four structures reveal conformational changes in prepore to pore conversion that support a multi-step mechanism by which low pH is sensed and the membrane-spanning channel is formed.

MeSH Terms
Antigens, Bacterial/chemistry,metabolism,ultrastructure Bacillus anthracis/chemistry,ultrastructure Bacterial Toxins/chemistry,metabolism Biocatalysis Cryoelectron Microscopy Hydrogen-Ion Concentration Ion Channels/chemistry,metabolism,ultrastructure Models, Molecular Phenylalanine/metabolism Protein Conformation Protein Transport Structure-Activity Relationship
Chemicals
Antigens, Bacterial Bacterial Toxins Ion Channels anthrax toxin Phenylalanine
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Jiang Jiansen
1] Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, California 90095, USA [2] California NanoSystems Institute, University of California, Los Angeles, California 90095, USA.
Pentelute Bradley L
Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Collier R John
Department of Microbiology and Immunobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Zhou Z Hong
1] Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, California 90095, USA [2] California NanoSystems Institute, University of California, Los Angeles, California 90095, USA.
References (47)
47 references, click to expand
  1. The protective antigen component of anthrax toxin forms functional octameric complexes.
    J Mol Biol. 2009 Sep 25;392(3):614-29 PMID: 19627991
  2. Structure of the yeast mitochondrial large ribosomal subunit.
    Science. 2014 Mar 28;343(6178):1485-9 PMID: 24675956
  3. Dominant-negative mutants of a toxin subunit: an approach to therapy of anthrax.
    Science. 2001 Apr 27;292(5517):695-7 PMID: 11326092
  4. How proteins adapt to a membrane-water interface.
    Trends Biochem Sci. 2000 Sep;25(9):429-34 PMID: 10973056
  5. MOLE 2.0: advanced approach for analysis of biomacromolecular channels.
    J Cheminform. 2013 Aug 16;5(1):39 PMID: 23953065
  6. EMAN: semiautomated software for high-resolution single-particle reconstructions.
    J Struct Biol. 1999 Dec 1;128(1):82-97 PMID: 10600563
  7. Anthrax toxin protective antigen integrates poly-γ-D-glutamate and pH signals to sense the optimal environment for channel formation.
    Proc Natl Acad Sci U S A. 2012 Nov 6;109(45):18378-83 PMID: 23100533
  8. Structure of heptameric protective antigen bound to an anthrax toxin receptor: a role for receptor in pH-dependent pore formation.
    Proc Natl Acad Sci U S A. 2004 Sep 7;101(36):13147-51 PMID: 15326297
  9. Accurate determination of local defocus and specimen tilt in electron microscopy.
    J Struct Biol. 2003 Jun;142(3):334-47 PMID: 12781660
  10. Crystal structure of a complex between anthrax toxin and its host cell receptor.
    Nature. 2004 Aug 19;430(7002):905-8 PMID: 15243628
  11. The geometry of the ribosomal polypeptide exit tunnel.
    J Mol Biol. 2006 Jul 21;360(4):893-906 PMID: 16784753
  12. Electrostatic ratchet in the protective antigen channel promotes anthrax toxin translocation.
    J Biol Chem. 2012 Dec 21;287(52):43753-64 PMID: 23115233
  13. Crystal structure of the Vibrio cholerae cytolysin heptamer reveals common features among disparate pore-forming toxins.
    Proc Natl Acad Sci U S A. 2011 May 3;108(18):7385-90 PMID: 21502531
  14. RELION: implementation of a Bayesian approach to cryo-EM structure determination.
    J Struct Biol. 2012 Dec;180(3):519-30 PMID: 23000701
  15. Charge requirements for proton gradient-driven translocation of anthrax toxin.
    J Biol Chem. 2011 Jul 1;286(26):23189-99 PMID: 21507946
  16. Anthrax toxin: receptor binding, internalization, pore formation, and translocation.
    Annu Rev Biochem. 2007;76:243-65 PMID: 17335404
  17. A phenylalanine clamp catalyzes protein translocation through the anthrax toxin pore.
    Science. 2005 Jul 29;309(5735):777-81 PMID: 16051798
  18. Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM.
    Nat Methods. 2013 Jun;10(6):584-90 PMID: 23644547
  19. Effects of introducing a single charged residue into the phenylalanine clamp of multimeric anthrax protective antigen.
    J Biol Chem. 2010 Mar 12;285(11):8130-7 PMID: 20061382
  20. UCSF Chimera--a visualization system for exploratory research and analysis.
    J Comput Chem. 2004 Oct;25(13):1605-12 PMID: 15264254
  21. Crystallography & NMR system: A new software suite for macromolecular structure determination.
    Acta Crystallogr D Biol Crystallogr. 1998 Sep 1;54(Pt 5):905-21 PMID: 9757107
  22. Bsoft: image and molecular processing in electron microscopy.
    J Struct Biol. 2001 Feb-Mar;133(2-3):156-69 PMID: 11472087
  23. Solubilization and characterization of the anthrax toxin pore in detergent micelles.
    Protein Sci. 2009 Sep;18(9):1882-95 PMID: 19609933
  24. Point mutations in anthrax protective antigen that block translocation.
    J Biol Chem. 2001 Mar 16;276(11):8371-6 PMID: 11113126
  25. Automated molecular microscopy: the new Leginon system.
    J Struct Biol. 2005 Jul;151(1):41-60 PMID: 15890530
  26. Proton-coupled protein transport through the anthrax toxin channel.
    Philos Trans R Soc Lond B Biol Sci. 2009 Jan 27;364(1514):209-15 PMID: 18957378
  27. Functions of phenylalanine residues within the beta-barrel stem of the anthrax toxin pore.
    PLoS One. 2009;4(7):e6280 PMID: 19609431
  28. Ribosome exit tunnel can entropically stabilize alpha-helices.
    Proc Natl Acad Sci U S A. 2005 Dec 27;102(52):18956-61 PMID: 16357202
  29. Visualization of substrate binding and translocation by the ATP-dependent protease, ClpXP.
    Mol Cell. 2000 Dec;6(6):1515-21 PMID: 11163224
  30. Structural and mechanistic insights into the bacterial amyloid secretion channel CsgG.
    Nature. 2014 Dec 11;516(7530):250-3 PMID: 25219853
  31. Structure of staphylococcal alpha-hemolysin, a heptameric transmembrane pore.
    Science. 1996 Dec 13;274(5294):1859-66 PMID: 8943190
  32. PHENIX: a comprehensive Python-based system for macromolecular structure solution.
    Acta Crystallogr D Biol Crystallogr. 2010 Feb;66(Pt 2):213-21 PMID: 20124702
  33. Resmap: automated representation of macromolecular interfaces as two-dimensional networks.
    Bioinformatics. 2005 Aug 1;21(15):3327-8 PMID: 15914544
  34. Binding stoichiometry and kinetics of the interaction of a human anthrax toxin receptor, CMG2, with protective antigen.
    J Biol Chem. 2004 May 28;279(22):23349-56 PMID: 15044490
  35. Fully automated, sequential tilt-series acquisition with Leginon.
    J Struct Biol. 2009 Jul;167(1):11-8 PMID: 19361558
  36. Ratcheting up protein translocation with anthrax toxin.
    Protein Sci. 2012 May;21(5):606-24 PMID: 22374876
  37. Trapping a translocating protein within the anthrax toxin channel: implications for the secondary structure of permeating proteins.
    J Gen Physiol. 2011 Apr;137(4):343-56 PMID: 21402886
  38. Protein translocation through the anthrax toxin transmembrane pore is driven by a proton gradient.
    J Mol Biol. 2006 Feb 3;355(5):968-79 PMID: 16343527
  39. Crystal structure of the anthrax toxin protective antigen.
    Nature. 1997 Feb 27;385(6619):833-8 PMID: 9039918
  40. Evidence for a proton-protein symport mechanism in the anthrax toxin channel.
    J Gen Physiol. 2009 Mar;133(3):307-14 PMID: 19204186
  41. Inference of macromolecular assemblies from crystalline state.
    J Mol Biol. 2007 Sep 21;372(3):774-97 PMID: 17681537
  42. Coot: model-building tools for molecular graphics.
    Acta Crystallogr D Biol Crystallogr. 2004 Dec;60(Pt 12 Pt 1):2126-32 PMID: 15572765
  43. Mapping dominant-negative mutations of anthrax protective antigen by scanning mutagenesis.
    Proc Natl Acad Sci U S A. 2003 Nov 25;100(24):13803-8 PMID: 14623961
  44. A Bayesian view on cryo-EM structure determination.
    J Mol Biol. 2012 Jan 13;415(2):406-18 PMID: 22100448
  45. Beam-induced motion correction for sub-megadalton cryo-EM particles.
    Elife. 2014;3:e03665 PMID: 25122622
  46. Structural basis for the unfolding of anthrax lethal factor by protective antigen oligomers.
    Nat Struct Mol Biol. 2010 Nov;17(11):1383-90 PMID: 21037566
  47. A loop network within the anthrax toxin pore positions the phenylalanine clamp in an active conformation.
    Proc Natl Acad Sci U S A. 2006 Jun 27;103(26):9802-7 PMID: 16785422
Article Info
Journal
Nature
Abbr.
Nature
ISSN
1476-4687
DOI
10.1038/nature14247
Published
2015-05-28
Epub
2015-00-16
Pages
545-9
Language
English
Region
England
NLM ID
0410462
PMCID
PMC4519040
Subset
IM
Grants
NCRR NIH HHS · 1S10RR23057 · United States
NIAID NIH HHS · R01 AI046420 · United States
NIAID NIH HHS · AI022021 · United States
NIH HHS · 1S10OD018111 · United States
NIAID NIH HHS · R01 AI094386 · United States
NIGMS NIH HHS · GM071940 · United States
NIH HHS · S10 OD018111 · United States
NIAID NIH HHS · AI046420 · United States
NIGMS NIH HHS · R01 GM071940 · United States
NIAID NIH HHS · AI094386 · United States
NIAID NIH HHS · AI057159 · United States
NCRR NIH HHS · S10 RR023057 · United States
Databases
PDB
3J9C
External Links
PubMed source DOI: 10.1038/nature14247 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2