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- EMDB-41282: CryoEM structure of octamer assembly of Shedu nuclease domain fro... -

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Basic information

Entry
Database: EMDB / ID: EMD-41282
TitleCryoEM structure of octamer assembly of Shedu nuclease domain from Bacillus cereus
Map dataCryoEM structure of Shedu from Bacillus cereus
Sample
  • Complex: Bacillus cereus Shedu delta_NL octamer
    • Protein or peptide: Shedu protein SduA
KeywordsShedu / DUF4263 / Bacterial defense systems / Nuclease / Anti-plasmid defense system / PD-(D/E)XK nuclease / Whirly domain / Two-component signaling / DNA BINDING PROTEIN
Function / homologyProtein of unknown function DUF4263 / : / Shedu protein SduA, C-terminal / Shedu protein SduA, N-terminal / nuclease activity / defense response to virus / Shedu protein SduA
Function and homology information
Biological speciesBacillus cereus B4264 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.19 Å
AuthorsGu Y / Corbett K
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R35 GM144121 United States
CitationJournal: to be published
Title: Shedu anti-phage nucleases share a common enzymatic core regulated by diverse sensor domains
Authors: Gu Y / Corbett K
History
DepositionJul 19, 2023-
Header (metadata) releaseJul 31, 2024-
Map releaseJul 31, 2024-
UpdateJul 31, 2024-
Current statusJul 31, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41282.png

Downloads & links

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Map

FileDownload / File: emd_41282.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM structure of Shedu from Bacillus cereus
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.94 Å/pix.
x 360 pix.
= 336.6 Å		0.94 Å/pix.
x 360 pix.
= 336.6 Å		0.94 Å/pix.
x 360 pix.
= 336.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.935 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.1
Minimum - Maximum-0.3204289 - 0.6380602
Average (Standard dev.)-0.00008402094 (±0.012005598)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 336.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: focused local refinement, main map

Fileemd_41282_additional_1.map
Annotationfocused local refinement, main map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: focused local refinement, half A map

Fileemd_41282_additional_2.map
Annotationfocused local refinement, half A map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: focused local refinement, half B map

Fileemd_41282_additional_3.map
Annotationfocused local refinement, half B map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map 1

Fileemd_41282_half_map_1.map
AnnotationHalf Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map 2

Fileemd_41282_half_map_2.map
AnnotationHalf Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Bacillus cereus Shedu delta_NL octamer

EntireName: Bacillus cereus Shedu delta_NL octamer
Components
  • Complex: Bacillus cereus Shedu delta_NL octamer
    • Protein or peptide: Shedu protein SduA

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Supramolecule #1: Bacillus cereus Shedu delta_NL octamer

SupramoleculeName: Bacillus cereus Shedu delta_NL octamer / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Details: Octamer assembly of Bacillus cereus Shedu nuclease domain, which truncates the N-terminal and linker region. Glutamic acid 264 is mutated to Alanine.
Source (natural)Organism: Bacillus cereus B4264 (bacteria)
Molecular weightTheoretical: 216 KDa

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Macromolecule #1: Shedu protein SduA

MacromoleculeName: Shedu protein SduA / type: protein_or_peptide / ID: 1 / Number of copies: 8 / Enantiomer: LEVO
Source (natural)Organism: Bacillus cereus B4264 (bacteria)
Molecular weightTheoretical: 26.23676 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MKSSHHHHHH ENLYFQSNAK EQDLDQLNTL IGIANLKKVL SVWESNKLTN TSEKFWQSVL KENTWILSQI FSNPTVLIND EAYVGGKTV KNDSGKLVDF LYANPFSKDA VLIAIKTPST PLITPTEYRT GVYSAHKDLT GAVTQVLTYK TTLQREYQNI D YNNYRQGI ...String:
MKSSHHHHHH ENLYFQSNAK EQDLDQLNTL IGIANLKKVL SVWESNKLTN TSEKFWQSVL KENTWILSQI FSNPTVLIND EAYVGGKTV KNDSGKLVDF LYANPFSKDA VLIAIKTPST PLITPTEYRT GVYSAHKDLT GAVTQVLTYK TTLQREYQNI D YNNYRQGI KTDFDIITPC CVVIAGMFDT LTDTAHRHSF ELYRKELKNV TVITFDELFE RVKGLIKLLE G

UniProtKB: Shedu protein SduA

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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

Concentration0.5 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
500.0 mMNaClSodium Chloride
20.0 mMC8H18N2O4SHEPES
1.0 mMC4H10O2S2Dithiothreitol
5.0 mMMgCl2Magnesium Chloride

Details: Prepared using deionized water and filtered strelized.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsFreshly collected from size-exclusion column

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Detector mode: COUNTING / Average electron dose: 55.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.2 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.19 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.2.1) / Number images used: 138326
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8ti9:
CryoEM structure of octamer assembly of Shedu nuclease domain from Bacillus cereus

PDB-8tia:
CryoEM structure of locally-refined tetramer of Shedu nuclease domain from Bacillus cereus

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