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- EMDB-42965: CryoEM structure of AriA-Ocr complex -

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Basic information

Entry
Database: EMDB / ID: EMD-42965
TitleCryoEM structure of AriA-Ocr complex
Map dataMain map
Sample
  • Complex: AriA hexamer with two monomers of Ocr
    • Protein or peptide: AriA antitoxin
    • Protein or peptide: Protein Ocr
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE
KeywordsBacterial Defense system / Toxin-antitoxin system / AriAB / PARIS / Immune system
Function / homologyB-form DNA mimic Ocr / DNA mimic ocr / Protein Ocr / symbiont-mediated evasion of host restriction-modification system / symbiont-mediated suppression of host innate immune response / : / Protein Ocr
Function and homology information
Biological speciesEscherichia coli B185 (bacteria) / Escherichia phage T7 (virus)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.63 Å
AuthorsDeep A / Corbett KD
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM144121 United States
CitationJournal: Nature / Year: 2024
Title: Architecture and activation mechanism of the bacterial PARIS defence system.
Authors: Amar Deep / Qishan Liang / Eray Enustun / Joe Pogliano / Kevin D Corbett /
Abstract: Bacteria and their viruses (bacteriophages or phages) are engaged in an intense evolutionary arms race. While the mechanisms of many bacterial antiphage defence systems are known, how these systems ...Bacteria and their viruses (bacteriophages or phages) are engaged in an intense evolutionary arms race. While the mechanisms of many bacterial antiphage defence systems are known, how these systems avoid toxicity outside infection yet activate quickly after infection is less well understood. Here we show that the bacterial phage anti-restriction-induced system (PARIS) operates as a toxin-antitoxin system, in which the antitoxin AriA sequesters and inactivates the toxin AriB until triggered by the T7 phage counterdefence protein Ocr. Using cryo-electron microscopy, we show that AriA is related to SMC-family ATPases but assembles into a distinctive homohexameric complex through two oligomerization interfaces. In uninfected cells, the AriA hexamer binds to up to three monomers of AriB, maintaining them in an inactive state. After Ocr binding, the AriA hexamer undergoes a structural rearrangement, releasing AriB and allowing it to dimerize and activate. AriB is a toprim/OLD-family nuclease, the activation of which arrests cell growth and inhibits phage propagation by globally inhibiting protein translation through specific cleavage of a lysine tRNA. Collectively, our findings reveal the intricate molecular mechanisms of a bacterial defence system triggered by a phage counterdefence protein, and highlight how an SMC-family ATPase has been adapted as a bacterial infection sensor.
History
DepositionNov 28, 2023-
Header (metadata) releaseJun 26, 2024-
Map releaseJun 26, 2024-
UpdateOct 16, 2024-
Current statusOct 16, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_42965.png

Downloads & links

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Map

FileDownload / File: emd_42965.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMain map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.94 Å/pix.
x 320 pix.
= 299.2 Å		0.94 Å/pix.
x 320 pix.
= 299.2 Å		0.94 Å/pix.
x 320 pix.
= 299.2 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: 0.07
Minimum - Maximum-0.21050641 - 0.43967268
Average (Standard dev.)0.00065672153 (±0.014455472)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 299.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Local Refinement map

Fileemd_42965_additional_1.map
AnnotationLocal Refinement map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_42965_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_42965_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : AriA hexamer with two monomers of Ocr

EntireName: AriA hexamer with two monomers of Ocr
Components
  • Complex: AriA hexamer with two monomers of Ocr
    • Protein or peptide: AriA antitoxin
    • Protein or peptide: Protein Ocr
  • Ligand: ADENOSINE-5'-TRIPHOSPHATE

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Supramolecule #1: AriA hexamer with two monomers of Ocr

SupramoleculeName: AriA hexamer with two monomers of Ocr / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2 / Details: AriA(6)-Ocr(2)
Source (natural)Organism: Escherichia coli B185 (bacteria)

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Macromolecule #1: AriA antitoxin

MacromoleculeName: AriA antitoxin / type: protein_or_peptide / ID: 1 / Number of copies: 6 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli B185 (bacteria)
Molecular weightTheoretical: 53.288098 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: VAIRTISKIE LSKIHNRYNL TVDFFNDLNV IHGKNGAGKS TLIHVIANIV NGDFIRFAFL IFEEIKATYS DGLKIVIRRD KIDEQSFIS VTLSNGKYIK FAVGEAMATV REIESERHLR ERDVKSMLAM DIDKFVKENE LQKVRASYFP AFRTMLEAWS S SSDVGYER ...String:
VAIRTISKIE LSKIHNRYNL TVDFFNDLNV IHGKNGAGKS TLIHVIANIV NGDFIRFAFL IFEEIKATYS DGLKIVIRRD KIDEQSFIS VTLSNGKYIK FAVGEAMATV REIESERHLR ERDVKSMLAM DIDKFVKENE LQKVRASYFP AFRTMLEAWS S SSDVGYER RVIRSSFYNR KASAFARELF GQFLPSINYP SPMEIEDRLR EEIRRAQLGI AAYESRTFSE SFVKVFSALF DN SSVEGEI TGELLKEIEG LAIAQDSSIK NGYYAEYSKV YEEIRSLINR NLKGKVENSV SGALVVYRDA LRDRQDYQEK AFS EIDNYM SSVNSFLEDK EMAYDFDLRR KYPKVGLKFP DGSWSPIRVL SSGERQLLTM LYAASKMGDD AIVLIDEPEI SLHI DWQED LLKRMLSQLS GRQIIVCTHS PSIATGYEDF MINISPEFIS SRDNDNHKDS EEMEEDESL

UniProtKB: UNIPROTKB: D6IC77

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Macromolecule #2: Protein Ocr

MacromoleculeName: Protein Ocr / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Escherichia phage T7 (virus)
Molecular weightTheoretical: 13.819015 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
MAMSNMTYNN VFDHAYEMLK ENIRYDDIRD TDDLHDAIHM AADNAVPHYY ADIFSVMASE GIDLEFEDSG LMPDTKDVIR ILQARIYEQ LTIDLWEDAE DLLNEYLEEV EEYEEDEE

UniProtKB: Protein Ocr

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Macromolecule #3: ADENOSINE-5'-TRIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-TRIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 4 / Formula: ATP
Molecular weightTheoretical: 507.181 Da
Chemical component information

ChemComp-ATP:
ADENOSINE-5'-TRIPHOSPHATE / ATP, energy-carrying molecule*YM

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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

Concentration2 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
150.0 mMKClPotassium chloride
20.0 mMTristris(hydroxymethyl)aminomethane
10.0 uMATPAdenosine triphosphate
2.0 mMBME2-Mercaptoethanol
10.0 mMMgCl2Magnesium chloride
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 12 sec. / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 278 K / Instrument: FEI VITROBOT MARK IV
DetailsSample was a mixture of three proteins.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris X / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Number grids imaged: 1 / Number real images: 6821 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionAlgorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.63 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 99697
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8v45:
CryoEM structure of AriA-Ocr complex

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