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- EMDB-13947: Cryo-EM structure of Botulinum neurotoxin serotype B -

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Basic information

Entry
Database: EMDB / ID: EMD-13947
TitleCryo-EM structure of Botulinum neurotoxin serotype B
Map data
Sample
  • Complex: Botulinum neurotoxin serotype B
    • Protein or peptide: Botulinum neurotoxin type B
Function / homology
Function and homology information


Toxicity of botulinum toxin type B (botB) / bontoxilysin / host cell presynaptic membrane / host cell cytoplasmic vesicle / host cell cytosol / protein transmembrane transporter activity / metalloendopeptidase activity / toxin activity / lipid binding / host cell plasma membrane ...Toxicity of botulinum toxin type B (botB) / bontoxilysin / host cell presynaptic membrane / host cell cytoplasmic vesicle / host cell cytosol / protein transmembrane transporter activity / metalloendopeptidase activity / toxin activity / lipid binding / host cell plasma membrane / proteolysis / zinc ion binding / extracellular region / membrane
Similarity search - Function
Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily ...Clostridium neurotoxin, translocation / Clostridium neurotoxin, Translocation domain / Clostridium neurotoxin, translocation domain / Clostridial neurotoxin zinc protease / Botulinum/Tetanus toxin, catalytic chain / Clostridium neurotoxin, receptor-binding C-terminal / Clostridium neurotoxin, receptor binding N-terminal / Clostridium neurotoxin, C-terminal receptor binding / Clostridium neurotoxin, N-terminal receptor binding / Kunitz inhibitor STI-like superfamily / Neutral zinc metallopeptidases, zinc-binding region signature. / Concanavalin A-like lectin/glucanase domain superfamily
Similarity search - Domain/homology
Botulinum neurotoxin type B
Similarity search - Component
Biological speciesClostridium botulinum (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsKosenina S / Martinez-Carranza M / Davies JR / Masuyer G / Stenmark P
Funding support Sweden, 2 items
OrganizationGrant numberCountry
Swedish Research Council2018-03406 Sweden
Cancerfonden20 1287 PjF Sweden
CitationJournal: Toxins (Basel) / Year: 2021
Title: Structural Analysis of Botulinum Neurotoxins Type B and E by Cryo-EM.
Authors: Sara Košenina / Markel Martínez-Carranza / Jonathan R Davies / Geoffrey Masuyer / Pål Stenmark /
Abstract: Botulinum neurotoxins (BoNTs) are the causative agents of a potentially lethal paralytic disease targeting cholinergic nerve terminals. Multiple BoNT serotypes exist, with types A, B and E being the ...Botulinum neurotoxins (BoNTs) are the causative agents of a potentially lethal paralytic disease targeting cholinergic nerve terminals. Multiple BoNT serotypes exist, with types A, B and E being the main cause of human botulism. Their extreme toxicity has been exploited for cosmetic and therapeutic uses to treat a wide range of neuromuscular disorders. Although naturally occurring BoNT types share a common end effect, their activity varies significantly based on the neuronal cell-surface receptors and intracellular SNARE substrates they target. These properties are the result of structural variations that have traditionally been studied using biophysical methods such as X-ray crystallography. Here, we determined the first structures of botulinum neurotoxins using single-particle cryogenic electron microscopy. The maps obtained at 3.6 and 3.7 Å for BoNT/B and /E, respectively, highlight the subtle structural dynamism between domains, and of the binding domain in particular. This study demonstrates how the recent advances made in the field of single-particle electron microscopy can be applied to bacterial toxins of clinical relevance and the botulinum neurotoxin family in particular.
History
DepositionDec 6, 2021-
Header (metadata) releaseJan 26, 2022-
Map releaseJan 26, 2022-
UpdateFeb 2, 2022-
Current statusFeb 2, 2022Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7qfq
  • Surface level: 0.2
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_13947.png

Downloads & links

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Map

FileDownload / File: emd_13947.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 300 pix.
= 327. Å		1.09 Å/pix.
x 300 pix.
= 327. Å		1.09 Å/pix.
x 300 pix.
= 327. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.09 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.252 / Movie #1: 0.2
Minimum - Maximum-1.6968719 - 2.180725
Average (Standard dev.)0.000328086 (±0.034015007)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 327.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.091.091.09
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z327.000327.000327.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-1.6972.1810.000

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Supplemental data

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Mask #1

Fileemd_13947_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: locally filtered map

Fileemd_13947_additional_1.map
Annotationlocally filtered map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: halp map A

Fileemd_13947_half_map_1.map
Annotationhalp map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map B

Fileemd_13947_half_map_2.map
Annotationhalf map B
Projections & Slices
AxesZYX

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Slices (1/2)
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Histogram

Histogram (log scale)

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Sample components

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Entire : Botulinum neurotoxin serotype B

EntireName: Botulinum neurotoxin serotype B
Components
  • Complex: Botulinum neurotoxin serotype B
    • Protein or peptide: Botulinum neurotoxin type B

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Supramolecule #1: Botulinum neurotoxin serotype B

SupramoleculeName: Botulinum neurotoxin serotype B / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Clostridium botulinum (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
Molecular weightTheoretical: 150 KDa

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Macromolecule #1: Botulinum neurotoxin type B

MacromoleculeName: Botulinum neurotoxin type B / type: protein_or_peptide / ID: 1
Details: catalytically inactive variant of botulinum neurotoxin serotype B [E231Q/H234Y]
Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Clostridium botulinum (bacteria)
Molecular weightTheoretical: 153.021922 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MPVTINNFNY NDPIDNNNII MMEPPFARGT GRYYKAFKIT DRIWIIPERY TFGYKPEDFN KSSGIFNRDV CEYYDPDYLN TNDKKNIFL QTMIKLFNRI KSKPLGEKLL EMIINGIPYL GDRRVPLEEF NTNIASVTVN KLISNPGEVE RKKGIFANLI I FGPGPVLN ...String:
MPVTINNFNY NDPIDNNNII MMEPPFARGT GRYYKAFKIT DRIWIIPERY TFGYKPEDFN KSSGIFNRDV CEYYDPDYLN TNDKKNIFL QTMIKLFNRI KSKPLGEKLL EMIINGIPYL GDRRVPLEEF NTNIASVTVN KLISNPGEVE RKKGIFANLI I FGPGPVLN ENETIDIGIQ NHFASREGFG GIMQMKFCPE YVSVFNNVQE NKGASIFNRR GYFSDPALIL MHQLIYVLHG LY GIKVDDL PIVPNEKKFF MQSTDAIQAE ELYTFGGQDP SIITPSTDKS IYDKVLQNFR GIVDRLNKVL VCISDPNINI NIY KNKFKD KYKFVEDSEG KYSIDVESFD KLYKSLMFGF TETNIAENYK IKTRASYFSD SLPPVKIKNL LDNEIYTIEE GFNI SDKDM EKEYRGQNKA INKQAYEEIS KEHLAVYKIQ MCKSVKAPGI CIDVDNEDLF FIADKNSFSD DLSKNERIEY NTQSN YIEN DFPINELILD TDLISKIELP SENTESLTDF NVDVPVYEKQ PAIKKIFTDE NTIFQYLYSQ TFPLDIRDIS LTSSFD DAL LFSNKVYSFF SMDYIKTANK VVEAGLFAGW VKQIVNDFVI EANKSNTMDK IADISLIVPY IGLALNVGNE TAKGNFE NA FEIAGASILL EFIPELLIPV VGAFLLESYI DNKNKIIKTI DNALTKRNEK WSDMYGLIVA QWLSTVNTQF YTIKEGMY K ALNYQAQALE EIIKYRYNIY SEKEKSNINI DFNDINSKLN EGINQAIDNI NNFINGCSVS YLMKKMIPLA VEKLLDFDN TLKKNLLNYI DENKLYLIGS AEYEKSKVNK YLKTIMPFDL SIYTNDTILI EMFNKYNSEI LNNIILNLRY KDNNLIDLSG YGAKVEVYD GVELNDKNQF KLTSSANSKI RVTQNQNIIF NSVFLDFSVS FWIRIPKYKN DGIQNYIHNE YTIINCMKNN S GWKISIRG NRIIWTLIDI NGKTKSVFFE YNIREDISEY INRWFFVTIT NNLNNAKIYI NGKLESNTDI KDIREVIANG EI IFKLDGD IDRTQFIWMK YFSIFNTELS QSNIEERYKI QSYSEYLKDF WGNPLMYNKE YYMFNAGNKN SYIKLKKDSP VGE ILTRSK YNQNSKYINY RDLYIGEKFI IRRKSNSQSI NDDIVRKEDY IYLDFFNLNQ EWRVYTYKYF KKEEMKLFLA PIYD SDEFY NTIQIKEYDE QPTYSCQLLF KKDEESTDEI GLIGIHRFYE SGIVFEEYKD YFCISKWYLK EVKRKPYNLK LGCNW QFIP KDEGWTELEV LFQGPLEHHH HHH

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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

Concentration0.05 mg/mL
BufferpH: 7.5 / Details: 20 mM HEPES pH7.5, 50mM NaCl, 0.5mM TCEP
GridModel: Quantifoil R0.6/1 / Material: COPPER / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV
DetailsPure protein, monodisperse by SEC

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: FEI FALCON III (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 2 / Number real images: 6317 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 2432309
CTF correctionSoftware - Name: cryoSPARC (ver. 3.1)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.1) / Number images used: 286802
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.1)
Final 3D classificationNumber classes: 19 / Software - Name: cryoSPARC (ver. 3.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementProtocol: FLEXIBLE FIT
Output model

PDB-7qfq:
Cryo-EM structure of Botulinum neurotoxin serotype B

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