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- EMDB-21898: CryoEM structure of the C. sordellii lethal toxin TcsL in complex... -

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Basic information

Entry
Database: EMDB / ID: EMD-21898
TitleCryoEM structure of the C. sordellii lethal toxin TcsL in complex with SEMA6A
Map data
Sample
  • Complex: Complex of SEMA6A with TcsL
    • Complex: SEMA6A
      • Protein or peptide: SEMA6A
    • Complex: TcsL
      • Protein or peptide: TcsL
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
KeywordscryoEM / signaling protein-toxin complex
Function / homology
Function and homology information


negative regulation of cell adhesion involved in sprouting angiogenesis / negative regulation of sprouting angiogenesis / semaphorin receptor binding / negative regulation of vascular endothelial growth factor signaling pathway / Other semaphorin interactions / positive regulation of neuron migration / chemorepellent activity / neural crest cell migration / host cell cytosol / Transferases; Glycosyltransferases; Hexosyltransferases ...negative regulation of cell adhesion involved in sprouting angiogenesis / negative regulation of sprouting angiogenesis / semaphorin receptor binding / negative regulation of vascular endothelial growth factor signaling pathway / Other semaphorin interactions / positive regulation of neuron migration / chemorepellent activity / neural crest cell migration / host cell cytosol / Transferases; Glycosyltransferases; Hexosyltransferases / glycosyltransferase activity / semaphorin-plexin signaling pathway / cellular response to vascular endothelial growth factor stimulus / cytoskeleton organization / cysteine-type peptidase activity / axon guidance / negative regulation of angiogenesis / host cell endosome membrane / animal organ morphogenesis / negative regulation of ERK1 and ERK2 cascade / nervous system development / toxin activity / cell surface receptor signaling pathway / axon / apoptotic process / host cell plasma membrane / extracellular region / metal ion binding / membrane / plasma membrane
Similarity search - Function
Semaphorin / TcdA/TcdB toxin, N-terminal helical domain / TcdB toxin N-terminal helical domain / TcdA/TcdB toxin, catalytic glycosyltransferase domain / TcdA/TcdB catalytic glycosyltransferase domain / TcdA/TcdB toxin, pore forming domain / TcdA/TcdB pore forming domain / CGT/MARTX, cysteine protease (CPD) domain / CGT/MARTX, cysteine protease (CPD) domain superfamily / Peptidase C80 family ...Semaphorin / TcdA/TcdB toxin, N-terminal helical domain / TcdB toxin N-terminal helical domain / TcdA/TcdB toxin, catalytic glycosyltransferase domain / TcdA/TcdB catalytic glycosyltransferase domain / TcdA/TcdB toxin, pore forming domain / TcdA/TcdB pore forming domain / CGT/MARTX, cysteine protease (CPD) domain / CGT/MARTX, cysteine protease (CPD) domain superfamily / Peptidase C80 family / CGT/MARTX cysteine protease (CPD) domain profile. / Choline-binding repeat / Putative cell wall binding repeat / Cell wall/choline-binding repeat / Cell wall-binding repeat profile. / Plexin repeat / Plexin repeat / Sema domain / semaphorin domain / Sema domain / Sema domain superfamily / Sema domain profile. / PSI domain / domain found in Plexins, Semaphorins and Integrins / Nucleotide-diphospho-sugar transferases / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Semaphorin-6A / Lethal Toxin
Similarity search - Component
Biological speciesHomo sapiens (human) / Paeniclostridium sordellii (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsKucharska I / Rubinstein JL
Funding support Canada, 3 items
OrganizationGrant numberCountry
Other governmentOntario Early Researcher Awards program Canada
Other governmentCanada Foundation for Innovation Canada
Other governmentOntario Research Fund Canada
CitationJournal: Cell / Year: 2020
Title: Recognition of Semaphorin Proteins by P. sordellii Lethal Toxin Reveals Principles of Receptor Specificity in Clostridial Toxins.
Authors: Hunsang Lee / Greg L Beilhartz / Iga Kucharska / Swetha Raman / Hong Cui / Mandy Hiu Yi Lam / Huazhu Liang / John L Rubinstein / Daniel Schramek / Jean-Philippe Julien / Roman A Melnyk / Mikko Taipale /
Abstract: Pathogenic clostridial species secrete potent toxins that induce severe host tissue damage. Paeniclostridium sordellii lethal toxin (TcsL) causes an almost invariably lethal toxic shock syndrome ...Pathogenic clostridial species secrete potent toxins that induce severe host tissue damage. Paeniclostridium sordellii lethal toxin (TcsL) causes an almost invariably lethal toxic shock syndrome associated with gynecological infections. TcsL is 87% similar to C. difficile TcdB, which enters host cells via Frizzled receptors in colon epithelium. However, P. sordellii infections target vascular endothelium, suggesting that TcsL exploits another receptor. Here, using CRISPR/Cas9 screening, we establish semaphorins SEMA6A and SEMA6B as TcsL receptors. We demonstrate that recombinant SEMA6A can protect mice from TcsL-induced edema. A 3.3 Å cryo-EM structure shows that TcsL binds SEMA6A with the same region that in TcdB binds structurally unrelated Frizzled. Remarkably, 15 mutations in this evolutionarily divergent surface are sufficient to switch binding specificity of TcsL to that of TcdB. Our findings establish semaphorins as physiologically relevant receptors for TcsL and reveal the molecular basis for the difference in tissue targeting and disease pathogenesis between highly related toxins.
History
DepositionMay 3, 2020-
Header (metadata) releaseJul 8, 2020-
Map releaseJul 8, 2020-
UpdateOct 9, 2024-
Current statusOct 9, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 2
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6wts
  • Surface level: 2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_21898.png

Downloads & links

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Map

FileDownload / File: emd_21898.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.03 Å/pix.
x 122 pix.
= 125.66 Å		1.03 Å/pix.
x 117 pix.
= 120.51 Å		1.03 Å/pix.
x 144 pix.
= 148.32 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 1.03 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.3 / Movie #1: 2
Minimum - Maximum-12.240899000000001 - 16.825357
Average (Standard dev.)0.000000000001972 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin866688
Dimensions117144122
Spacing122117144
CellA: 125.659996 Å / B: 120.509995 Å / C: 148.31999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.031.031.03
M x/y/z122117144
origin x/y/z0.0000.0000.000
length x/y/z125.660120.510148.320
α/β/γ90.00090.00090.000
start NX/NY/NZ888666
NX/NY/NZ122117144
MAP C/R/S321
start NC/NR/NS668688
NC/NR/NS144117122
D min/max/mean-12.24116.8250.000

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Supplemental data

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Sample components

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Entire : Complex of SEMA6A with TcsL

EntireName: Complex of SEMA6A with TcsL
Components
  • Complex: Complex of SEMA6A with TcsL
    • Complex: SEMA6A
      • Protein or peptide: SEMA6A
    • Complex: TcsL
      • Protein or peptide: TcsL
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: Complex of SEMA6A with TcsL

SupramoleculeName: Complex of SEMA6A with TcsL / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2

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Supramolecule #2: SEMA6A

SupramoleculeName: SEMA6A / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: TcsL

SupramoleculeName: TcsL / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Source (natural)Organism: Paeniclostridium sordellii (bacteria)

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Macromolecule #1: SEMA6A

MacromoleculeName: SEMA6A / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 63.85741 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: ETGFPEDSEP ISISHGNYTK QYPVFVGHKP GRNTTQRHRL DIQMIMIMNG TLYIAARDHI YTVDIDTSHT EEIYCSKKLT WKSRQADVD TCRMKGKHKD ECHNFIKVLL KKNDDALFVC GTNAFNPSCR NYKMDTLEPF GDEFSGMARC PYDAKHANVA L FADGKLYS ...String:
ETGFPEDSEP ISISHGNYTK QYPVFVGHKP GRNTTQRHRL DIQMIMIMNG TLYIAARDHI YTVDIDTSHT EEIYCSKKLT WKSRQADVD TCRMKGKHKD ECHNFIKVLL KKNDDALFVC GTNAFNPSCR NYKMDTLEPF GDEFSGMARC PYDAKHANVA L FADGKLYS ATVTDFLAID AVIYRSLGES PTLRTVKHDS KWLKEPYFVQ AVDYGDYIYF FFREIAVEYN TMGKVVFPRV AQ VCKNDMG GSQRVLEKQW TSFLKARLNC SVPGDSHFYF NILQAVTDVI RINGRDVVLA TFSTPYNSIP GSAVCAYDML DIA SVFTGR FKEQKSPDST WTPVPDERVP KPRPGCCAGS SSLERYATSN EFPDDTLNFI KTHPLMDEAV PSIFNRPWFL RTMV RYRLT KIAVDTAAGP YQNHTVVFLG SEKGIILKFL ARIGNSGFLN DSLFLEEMSV YNSEKCSYDG VEDKRIMGMQ LDRAS SSLY VAFSTCVIKV PLGRCERYGK CKKTCIASRD PYCGWIKEGG ACSHLSPNSR LTFEQDIERG NTDGLGDCHN GSWSHP QFE K

UniProtKB: Semaphorin-6A

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Macromolecule #2: TcsL

MacromoleculeName: TcsL / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
EC number: Transferases; Glycosyltransferases; Hexosyltransferases
Source (natural)Organism: Paeniclostridium sordellii (bacteria)
Molecular weightTheoretical: 63.315875 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: SAWSHPQFEK ENLYFQGTNV RINLDGNTRS FIVPVITTEQ IRKNLSYSFY GSGGSYSLSL SPYNMNIDLN LVENDTWVID VDNVVKNIT IESDEIQKGE LIENILSKLN IEDNKIVLNN HTINFYGAIN ESNRFISLTF SILEDINIII EIDLVSKSYK I LLSGNCIK ...String:
SAWSHPQFEK ENLYFQGTNV RINLDGNTRS FIVPVITTEQ IRKNLSYSFY GSGGSYSLSL SPYNMNIDLN LVENDTWVID VDNVVKNIT IESDEIQKGE LIENILSKLN IEDNKIVLNN HTINFYGAIN ESNRFISLTF SILEDINIII EIDLVSKSYK I LLSGNCIK LIENSSDIQQ KIDHIGFNGE HQKYIPYSYI DNETKYNGFI DYSKKEGLFT AEFSNESIIR NIYMPDSNNL FI YSSKDLK DIRIINKGDV KLLIGNYFKD NMKVSLSFTI EDTNTIKLNG VYLDENGVAQ ILKFMNNAKS ALNTSNSLMN FLE SINIKN IFYNNLDPNI KFILDTNFII SGSNSIGQFE LICDKDKNIQ PYFIKFKIKE TSYTLYAGNR QNLIVEPSYH LDDS GNISS TVINFSQKYL YGIDRYVNKV IITPNLYTDE INITPVYKPN YICPEVIILD ANYINEKINI NINDLSIRYV WDNDG SDLI LIANSEEDNQ PQVKIRFVNV FKSDTAADKL SFNFSDKQDV SVSKIISTFS LAAGSENLYF QGHHHHHH

UniProtKB: Lethal Toxin

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 4 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.9 mg/mL
BufferpH: 7
VitrificationCryogen name: ETHANE-PROPANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: OTHER / Average electron dose: 45.24 e/Å2 / Details: Dataset was collected with FEI Falcon IV.
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

6c0b

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 179188
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE

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Atomic model buiding 1

Initial modelPDB ID:

6c0b


Chain - Source name: PDB / Chain - Initial model type: experimental model
Output model

PDB-6wts:
CryoEM structure of the C. sordellii lethal toxin TcsL in complex with SEMA6A

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