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- EMDB-29325: Map of RdrA from Escherichia coli RADAR defense system in double-... -

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Basic information

Entry
Database: EMDB / ID: EMD-29325
TitleMap of RdrA from Escherichia coli RADAR defense system in double-split conformation
Map dataE. coli RdrA, double split map
Sample
  • Complex: E. coli RdrA in double-split conformation
KeywordsRADAR / anti-phage defense / ATPase / IMMUNE SYSTEM
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.35 Å
AuthorsDuncan-Lowey B / Johnson AG / Rawson S / Mayer ML / Kranzusch PJ
Funding support United States, 4 items
OrganizationGrant numberCountry
The Pew Charitable Trusts United States
Burroughs Wellcome Fund United States
The G. Harold and Leila Y. Mathers Foundation United States
Parker Institute for Cancer Immunotherapy United States
CitationJournal: Cell / Year: 2023
Title: Cryo-EM structure of the RADAR supramolecular anti-phage defense complex.
Authors: Brianna Duncan-Lowey / Nitzan Tal / Alex G Johnson / Shaun Rawson / Megan L Mayer / Shany Doron / Adi Millman / Sarah Melamed / Taya Fedorenko / Assaf Kacen / Alexander Brandis / Tevie ...Authors: Brianna Duncan-Lowey / Nitzan Tal / Alex G Johnson / Shaun Rawson / Megan L Mayer / Shany Doron / Adi Millman / Sarah Melamed / Taya Fedorenko / Assaf Kacen / Alexander Brandis / Tevie Mehlman / Gil Amitai / Rotem Sorek / Philip J Kranzusch /
Abstract: RADAR is a two-protein bacterial defense system that was reported to defend against phage by "editing" messenger RNA. Here, we determine cryo-EM structures of the RADAR defense complex, revealing ...RADAR is a two-protein bacterial defense system that was reported to defend against phage by "editing" messenger RNA. Here, we determine cryo-EM structures of the RADAR defense complex, revealing RdrA as a heptameric, two-layered AAA+ ATPase and RdrB as a dodecameric, hollow complex with twelve surface-exposed deaminase active sites. RdrA and RdrB join to form a giant assembly up to 10 MDa, with RdrA docked as a funnel over the RdrB active site. Surprisingly, our structures reveal an RdrB active site that targets mononucleotides. We show that RdrB catalyzes ATP-to-ITP conversion in vitro and induces the massive accumulation of inosine mononucleotides during phage infection in vivo, limiting phage replication. Our results define ATP mononucleotide deamination as a determinant of RADAR immunity and reveal supramolecular assembly of a nucleotide-modifying machine as a mechanism of anti-phage defense.
History
DepositionDec 28, 2022-
Header (metadata) releaseFeb 1, 2023-
Map releaseFeb 1, 2023-
UpdateJan 17, 2024-
Current statusJan 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_29325.png

Downloads & links

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Map

FileDownload / File: emd_29325.map.gz / Format: CCP4 / Size: 274.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationE. coli RdrA, double split map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 416 pix.
= 343.2 Å		0.83 Å/pix.
x 416 pix.
= 343.2 Å		0.83 Å/pix.
x 416 pix.
= 343.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.825 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.95
Minimum - Maximum-2.471181 - 4.469298
Average (Standard dev.)-0.0009168522 (±0.10983601)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions416416416
Spacing416416416
CellA=B=C: 343.19998 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: E. coli RdrA, double split half map A

Fileemd_29325_half_map_1.map
AnnotationE. coli RdrA, double split half map A
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: E. coli RdrA, double split half map B

Fileemd_29325_half_map_2.map
AnnotationE. coli RdrA, double split half map B
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : E. coli RdrA in double-split conformation

EntireName: E. coli RdrA in double-split conformation
Components
  • Complex: E. coli RdrA in double-split conformation

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Supramolecule #1: E. coli RdrA in double-split conformation

SupramoleculeName: E. coli RdrA in double-split conformation / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Escherichia coli (E. coli)

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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
Component:
ConcentrationFormulaName
100.0 mMKClpotassium chloride
50.0 mMHEPES
1.0 mMTCEP
VitrificationCryogen name: ETHANE / Chamber humidity: 100 %

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 48.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER / Nominal defocus max: 2.5 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: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.35 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 421521
Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER

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