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- EMDB-43105: Map of the AriA homohexamer following release of the AriB effecto... -

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Basic information

Entry
Database: EMDB / ID: EMD-43105
TitleMap of the AriA homohexamer following release of the AriB effector during PARIS-mediated defense.
Map dataSharpened map of the AriA homohexamer.
Sample
  • Complex: Map of AriA homohexamer obtained from pulldowns of PARIS-Ocr coexpression showing AriB dissociated from AriA upon activation of the PARIS defense system.
    • Protein or peptide: AriA, ABC ATPase and sensor of PARIS immunity.
KeywordsPARIS / AriA / AriB / DUF4435 / IMMUNE SYSTEM
Biological speciesEscherichia coli B185 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.5 Å
AuthorsBurman NB / Santiago-Frangos A / Henriques W / Wilkinson R / Graham A / Wiedenheft B
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM134867 United States
CitationJournal: Nature / Year: 2024
Title: A virally encoded tRNA neutralizes the PARIS antiviral defence system.
Authors: Nathaniel Burman / Svetlana Belukhina / Florence Depardieu / Royce A Wilkinson / Mikhail Skutel / Andrew Santiago-Frangos / Ava B Graham / Alexei Livenskyi / Anna Chechenina / Natalia ...Authors: Nathaniel Burman / Svetlana Belukhina / Florence Depardieu / Royce A Wilkinson / Mikhail Skutel / Andrew Santiago-Frangos / Ava B Graham / Alexei Livenskyi / Anna Chechenina / Natalia Morozova / Trevor Zahl / William S Henriques / Murat Buyukyoruk / Christophe Rouillon / Baptiste Saudemont / Lena Shyrokova / Tatsuaki Kurata / Vasili Hauryliuk / Konstantin Severinov / Justine Groseille / Agnès Thierry / Romain Koszul / Florian Tesson / Aude Bernheim / David Bikard / Blake Wiedenheft / Artem Isaev /
Abstract: Viruses compete with each other for limited cellular resources, and some deliver defence mechanisms that protect the host from competing genetic parasites. The phage antirestriction induced system ...Viruses compete with each other for limited cellular resources, and some deliver defence mechanisms that protect the host from competing genetic parasites. The phage antirestriction induced system (PARIS) is a defence system, often encoded in viral genomes, that is composed of a 55 kDa ABC ATPase (AriA) and a 35 kDa TOPRIM nuclease (AriB). However, the mechanism by which AriA and AriB function in phage defence is unknown. Here we show that AriA and AriB assemble into a 425 kDa supramolecular immune complex. We use cryo-electron microscopy to determine the structure of this complex, thereby explaining how six molecules of AriA assemble into a propeller-shaped scaffold that coordinates three subunits of AriB. ATP-dependent detection of foreign proteins triggers the release of AriB, which assembles into a homodimeric nuclease that blocks infection by cleaving host lysine transfer RNA. Phage T5 subverts PARIS immunity through expression of a lysine transfer RNA variant that is not cleaved by PARIS, thereby restoring viral infection. Collectively, these data explain how AriA functions as an ATP-dependent sensor that detects viral proteins and activates the AriB toxin. PARIS is one of an emerging set of immune systems that form macromolecular complexes for the recognition of foreign proteins, rather than foreign nucleic acids.
History
DepositionDec 11, 2023-
Header (metadata) releaseSep 25, 2024-
Map releaseSep 25, 2024-
UpdateApr 9, 2025-
Current statusApr 9, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43105.png

Downloads & links

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Map

FileDownload / File: emd_43105.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map of the AriA homohexamer.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 300 pix.
= 331.2 Å		1.1 Å/pix.
x 300 pix.
= 331.2 Å		1.1 Å/pix.
x 300 pix.
= 331.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.104 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05
Minimum - Maximum-0.0011770197 - 1.9583793
Average (Standard dev.)0.002441709 (±0.036172107)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 331.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map A used in sharpening and gold standard FSC calculation

Fileemd_43105_half_map_1.map
AnnotationHalf map A used in sharpening and gold standard FSC calculation
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map B used in sharpening and gold standard FSC calculation

Fileemd_43105_half_map_2.map
AnnotationHalf map B used in sharpening and gold standard FSC calculation
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Map of AriA homohexamer obtained from pulldowns of PARIS-Ocr coex...

EntireName: Map of AriA homohexamer obtained from pulldowns of PARIS-Ocr coexpression showing AriB dissociated from AriA upon activation of the PARIS defense system.
Components
  • Complex: Map of AriA homohexamer obtained from pulldowns of PARIS-Ocr coexpression showing AriB dissociated from AriA upon activation of the PARIS defense system.
    • Protein or peptide: AriA, ABC ATPase and sensor of PARIS immunity.

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Supramolecule #1: Map of AriA homohexamer obtained from pulldowns of PARIS-Ocr coex...

SupramoleculeName: Map of AriA homohexamer obtained from pulldowns of PARIS-Ocr coexpression showing AriB dissociated from AriA upon activation of the PARIS defense system.
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli B185 (bacteria)

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Macromolecule #1: AriA, ABC ATPase and sensor of PARIS immunity.

MacromoleculeName: AriA, ABC ATPase and sensor of PARIS immunity. / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Escherichia coli B185 (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAIRTISKIE LSKIHNRYNL TVDFFNDLNV IHGKNGAGKS TLIHVIANIV NGDFIRFAFL IFEEIKATYS DGLKIVIRRD KIDEQSFISV TLSNGKYIKF AVGEAMATVR EIESERHLRE RDVKSMLAMD IDKFVKENEL QKVRASYFPA FRTMLEAWSS SSDVGYERRV ...String:
MAIRTISKIE LSKIHNRYNL TVDFFNDLNV IHGKNGAGKS TLIHVIANIV NGDFIRFAFL IFEEIKATYS DGLKIVIRRD KIDEQSFISV TLSNGKYIKF AVGEAMATVR EIESERHLRE RDVKSMLAMD IDKFVKENEL QKVRASYFPA FRTMLEAWSS SSDVGYERRV IRSSFYNRKA SAFARELFGQ FLPSINYPSP MEIEDRLREE IRRAQLGIAA YESRTFSESF VKVFSALFDN SSVEGEITGE LLKEIEGLAI AQDSSIKNGY YAEYSKVYEE IRSLINRNLK GKVENSVSGA LVVYRDALRD RQDYQEKAFS EIDNYMSSVN SFLEDKEMAY DFDLRRKYPK VGLKFPDGSW SPIRVLSSGE RQLLTMLYAA SKMGDDAIVL IDEPEISLHI DWQEDLLKRM LSQLSGRQII VCTHSPSIAT GYEDFMINIS PEFISSRDND NHKDSEEMEE DESL

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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

BufferpH: 7.5
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 10340 / Average electron dose: 56.31 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 36000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 4415782 / Details: Particles Extracted for initial classification
Startup modelType of model: NONE
Details: Multiclass ab-initio reconstructions were used to sort 667,479 particle images from 7 selected 2-D classes and obtain a consensus refinement.
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C3 (3 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.41) / Software - details: Non-Uniform Refinement / Number images used: 62732
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.41) / Software - details: Ab-Initio Reconstruction
Details: Further particle sorting yielded a class of 392989 particles that were further sorted by an additional round of 2-D classification.
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.41) / Software - details: Non-Uniform Refinement
Final 3D classificationNumber classes: 50 / Avg.num./class: 9169 / Software - Name: cryoSPARC (ver. 4.41) / Software - details: Heterogenous Refinement
Details: From a classification of 264,881 particles, 11 classes containing 100,862 particles were kept for further processing. These 100,862 particles were input to a 2-class ab initio ...Details: From a classification of 264,881 particles, 11 classes containing 100,862 particles were kept for further processing. These 100,862 particles were input to a 2-class ab initio reconstruction, followed by a 2 class heterogenous refinement to yield the final stack of 62,732 particles for the final reconstruction.
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChainDetails

8ux9

chain_id: B, source_name: PDB, initial_model_type: experimental model8UX9 contains two molecules of AriA. This reconstruction lacks the AriB subunit and contains 3 copies of chain B from the asymmetric unit.

8ux9

chain_id: C, source_name: PDB, initial_model_type: experimental model8UX9 contains two molecules of AriA. This reconstruction lacks the AriB subunit and contains 3 copies of chain C from the asymmetric unit.
DetailsRigid body fitting was done using the fitmap command in ChimeraX to dock 3 copies of the experimentally determined structure of the asymmetric unit into the density map of the fully assembled complex.
RefinementSpace: RECIPROCAL / Protocol: RIGID BODY FIT / Overall B value: 158.6 / Target criteria: Cross-correlation coefficient

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