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- EMDB-11524: Fully-loaded anthrax lethal toxin in its heptameric pre-pore stat... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-11524 | |||||||||
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Title | Fully-loaded anthrax lethal toxin in its heptameric pre-pore state and PA7LF(2+1A) arrangement | |||||||||
![]() | PA7LF(2 1A) | |||||||||
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![]() | anthrax lethal toxin / fully-loaded pre-pore state / membrane translocase / cytotoxic substrate / TOXIN | |||||||||
Function / homology | ![]() anthrax lethal factor endopeptidase / positive regulation of apoptotic process in another organism / host cell cytosol / negative regulation of MAPK cascade / Uptake and function of anthrax toxins / host cell endosome membrane / protein homooligomerization / metalloendopeptidase activity / metallopeptidase activity / toxin activity ...anthrax lethal factor endopeptidase / positive regulation of apoptotic process in another organism / host cell cytosol / negative regulation of MAPK cascade / Uptake and function of anthrax toxins / host cell endosome membrane / protein homooligomerization / metalloendopeptidase activity / metallopeptidase activity / toxin activity / host cell plasma membrane / proteolysis / zinc ion binding / extracellular region / identical protein binding / membrane / metal ion binding Similarity search - Function | |||||||||
Biological species | ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.2 Å | |||||||||
![]() | Quentin D / Antoni C | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Cryo-EM structure of the fully-loaded asymmetric anthrax lethal toxin in its heptameric pre-pore state. Authors: Claudia Antoni / Dennis Quentin / Alexander E Lang / Klaus Aktories / Christos Gatsogiannis / Stefan Raunser / ![]() Abstract: Anthrax toxin is the major virulence factor secreted by Bacillus anthracis, causing high mortality in humans and other mammals. It consists of a membrane translocase, known as protective antigen (PA) ...Anthrax toxin is the major virulence factor secreted by Bacillus anthracis, causing high mortality in humans and other mammals. It consists of a membrane translocase, known as protective antigen (PA), that catalyzes the unfolding of its cytotoxic substrates lethal factor (LF) and edema factor (EF), followed by translocation into the host cell. Substrate recruitment to the heptameric PA pre-pore and subsequent translocation, however, are not well understood. Here, we report three high-resolution cryo-EM structures of the fully-loaded anthrax lethal toxin in its heptameric pre-pore state, which differ in the position and conformation of LFs. The structures reveal that three LFs interact with the heptameric PA and upon binding change their conformation to form a continuous chain of head-to-tail interactions. As a result of the underlying symmetry mismatch, one LF binding site in PA remains unoccupied. Whereas one LF directly interacts with a part of PA called α-clamp, the others do not interact with this region, indicating an intermediate state between toxin assembly and translocation. Interestingly, the interaction of the N-terminal domain with the α-clamp correlates with a higher flexibility in the C-terminal domain of the protein. Based on our data, we propose a model for toxin assembly, in which the relative position of the N-terminal α-helices in the three LFs determines which factor is translocated first. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 137.7 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 18.5 KB 18.5 KB | Display Display | ![]() |
Images | ![]() | 161.9 KB | ||
Filedesc metadata | ![]() | 7.5 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6zxlMC ![]() 6zxjC ![]() 6zxkC M: atomic model generated by this map C: citing same article ( |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
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Annotation | PA7LF(2 1A) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.07 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
-Entire : Fully-loaded anthrax lethal toxin in its heptameric pre-pore stat...
Entire | Name: Fully-loaded anthrax lethal toxin in its heptameric pre-pore state and PA7LF(2+1A) arrangement |
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Components |
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-Supramolecule #1: Fully-loaded anthrax lethal toxin in its heptameric pre-pore stat...
Supramolecule | Name: Fully-loaded anthrax lethal toxin in its heptameric pre-pore state and PA7LF(2+1A) arrangement type: complex / ID: 1 / Parent: 0 / Macromolecule list: all Details: Heptameric pre-pores of proteolytically-acitivated protective antigen were loaded with excess of LFs to create the PA7LF3 complexes. The LFs form a continuous chain of head-to-tail ...Details: Heptameric pre-pores of proteolytically-acitivated protective antigen were loaded with excess of LFs to create the PA7LF3 complexes. The LFs form a continuous chain of head-to-tail interactions. In the PA7LF(2+1A) arrangement, the third LF binds with its N-terminal domain to the C-terminal region of the 2nd LF. |
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Molecular weight | Theoretical: 93 KDa |
-Supramolecule #2: Protective antigen
Supramolecule | Name: Protective antigen / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1 Details: The trypsin-activated 63 kDa fragments assemble into a hepatameric pre-pore |
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Source (natural) | Organism: ![]() ![]() |
-Supramolecule #3: Lethal factor
Supramolecule | Name: Lethal factor / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2 Details: Three LF molecules crown the heptameric PA ring. The LFs form a continuous chain of head-to-tail interactions. In the PA7LF(2+1A) arrangement, the third LF binds with its N-terminal domain ...Details: Three LF molecules crown the heptameric PA ring. The LFs form a continuous chain of head-to-tail interactions. In the PA7LF(2+1A) arrangement, the third LF binds with its N-terminal domain to the C-terminus of the 2nd LF. |
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Source (natural) | Organism: ![]() ![]() |
-Macromolecule #1: Protective antigen
Macromolecule | Name: Protective antigen / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 85.67993 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MGHHHHHHHH HHSSGHIDDD DKHMEVKQEN RLLNESESSS QGLLGYYFSD LNFQAPMVVT SSTTGDLSIP SSELENIPSE NQYFQSAIW SGFIKVKKSD EYTFATSADN HVTMWVDDQE VINKASNSNK IRLEKGRLYQ IKIQYQRENP TEKGLDFKLY W TDSQNKKE ...String: MGHHHHHHHH HHSSGHIDDD DKHMEVKQEN RLLNESESSS QGLLGYYFSD LNFQAPMVVT SSTTGDLSIP SSELENIPSE NQYFQSAIW SGFIKVKKSD EYTFATSADN HVTMWVDDQE VINKASNSNK IRLEKGRLYQ IKIQYQRENP TEKGLDFKLY W TDSQNKKE VISSDNLQLP ELKQKSSNSR KKRSTSAGPT VPDRDNDGIP DSLEVEGYTV DVKNKRTFLS PWISNIHEKK GL TKYKSSP EKWSTASDPY SDFEKVTGRI DKNVSPEARH PLVAAYPIVH VDMENIILSK NEDQSTQNTD SQTRTISKNT STS RTHTSE VHGNAEVHAS FFDIGGSVSA GFSNSNSSTV AIDHSLSLAG ERTWAETMGL NTADTARLNA NIRYVNTGTA PIYN VLPTT SLVLGKNQTL ATIKAKENQL SQILAPNNYY PSKNLAPIAL NAQDDFSSTP ITMNYNQFLE LEKTKQLRLD TDQVY GNIA TYNFENGRVR VDTGSNWSEV LPQIQETTAR IIFNGKDLNL VERRIAAVNP SDPLETTKPD MTLKEALKIA FGFNEP NGN LQYQGKDITE FDFNFDQQTS QNIKNQLAEL NATNIYTVLD KIKLNAKMNI LIRDKRFHYD RNNIAVGADE SVVKEAH RE VINSSTEGLL LNIDKDIRKI LSGYIVEIED TEGLKEVIND RYDMLNISSL RQDGKTFIDF KKYNDKLPLY ISNPNYKV N VYAVTKENTI INPSENGDTS TNGIKKILIF SKKGYEIG UniProtKB: Protective antigen |
-Macromolecule #2: Lethal factor
Macromolecule | Name: Lethal factor / type: protein_or_peptide / ID: 2 / Number of copies: 3 / Enantiomer: LEVO / EC number: anthrax lethal factor endopeptidase |
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Source (natural) | Organism: ![]() ![]() |
Molecular weight | Theoretical: 93.904211 KDa |
Recombinant expression | Organism: ![]() ![]() |
Sequence | String: MNIKKEFIKV ISMSCLVTAI TLSGPVFIPL VQGAGGHGDV GMHVKEKEKN KDENKRKDEE RNKTQEEHLK EIMKHIVKIE VKGEEAVKK EAAEKLLEKV PSDVLEMYKA IGGKIYIVDG DITKHISLEA LSEDKKKIKD IYGKDALLHE HYVYAKEGYE P VLVIQSSE ...String: MNIKKEFIKV ISMSCLVTAI TLSGPVFIPL VQGAGGHGDV GMHVKEKEKN KDENKRKDEE RNKTQEEHLK EIMKHIVKIE VKGEEAVKK EAAEKLLEKV PSDVLEMYKA IGGKIYIVDG DITKHISLEA LSEDKKKIKD IYGKDALLHE HYVYAKEGYE P VLVIQSSE DYVENTEKAL NVYYEIGKIL SRDILSKINQ PYQKFLDVLN TIKNASDSDG QDLLFTNQLK EHPTDFSVEF LE QNSNEVQ EVFAKAFAYY IEPQHRDVLQ LYAPEAFNYM DKFNEQEINL SLEELKDQRM LARYEKWEKI KQHYQHWSDS LSE EGRGLL KKLQIPIEPK KDDIIHSLSQ EEKELLKRIQ IDSSDFLSTE EKEFLKKLQI DIRDSLSEEE KELLNRIQVD SSNP LSEKE KEFLKKLKLD IQPYDINQRL QDTGGLIDSP SINLDVRKQY KRDIQNIDAL LHQSIGSTLY NKIYLYENMN INNLT ATLG ADLVDSTDNT KINRGIFNEF KKNFKYSISS NYMIVDINER PALDNERLKW RIQLSPDTRA GYLENGKLIL QRNIGL EIK DVQIIKQSEK EYIRIDAKVV PKSKIDTKIQ EAQLNINQEW NKALGLPKYT KLITFNVHNR YASNIVESAY LILNEWK NN IQSDLIKKVT NYLVDGNGRF VFTDITLPNI AEQYTHQDEI YEQVHSKGLY VPESRSILLH GPSKGVELRN DSEGFIHE F GHAVDDYAGY LLDKNQSDLV TNSKKFIDIF KEEGSNLTSY GRTNEAEFFA EAFRLMHSTD HAERLKVQKN APKTFQFIN DQIKFIINS UniProtKB: Lethal factor |
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 0.06 mg/mL | |||||||||
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Buffer | pH: 8.5 Component:
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Grid | Model: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec. / Pretreatment - Atmosphere: AIR | |||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 286 K / Instrument: GATAN CRYOPLUNGE 3 Details: 4 uL sample was applied to grid (with 2 nm additional carbon layer) and incubated for 45 s prior blotting.. |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number real images: 5238 / Average exposure time: 15.0 sec. / Average electron dose: 74.4 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | Calibrated defocus max: 2.6 µm / Calibrated defocus min: 1.2 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal magnification: 130000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Details | The previously generated PA7LF(2+1B) model (PDB:6ZXK) served as starting point and was placed into the density using rigid-body fit in Chimera. From this model, chain J was fitted into the density corresponding to the third LF, located adjacent to the second LF. For the entire model a restrained refinement in phenix was performed. The resulting model was further refined with a combination of phenix and coot. Unresolved regions were deleted and side chain information was removed for less well-resolved regions. |
Output model | ![]() PDB-6zxl: |