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- EMDB-29326: Structure of RdrA from Streptococcus suis RADAR defense system -

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Basic information

Entry
Database: EMDB / ID: EMD-29326
TitleStructure of RdrA from Streptococcus suis RADAR defense system
Map data
Sample
  • Complex: RdrA heptameric complex
    • Protein or peptide: KAP NTPase domain-containing protein
Function / homologyKAP family P-loop domain / KAP family P-loop domain / P-loop containing nucleoside triphosphate hydrolase / KAP NTPase domain-containing protein
Function and homology information
Biological speciesStreptococcus suis (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.5 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-
UpdateMar 15, 2023-
Current statusMar 15, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_29326.png

Downloads & links

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Map

FileDownload / File: emd_29326.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.1 Å
Density
Contour LevelBy AUTHOR: 1.6
Minimum - Maximum-7.2222695 - 12.103292
Average (Standard dev.)-0.00072370394 (±0.26373768)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 440.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_29326_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_29326_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 : RdrA heptameric complex

EntireName: RdrA heptameric complex
Components
  • Complex: RdrA heptameric complex
    • Protein or peptide: KAP NTPase domain-containing protein

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Supramolecule #1: RdrA heptameric complex

SupramoleculeName: RdrA heptameric complex / type: complex / ID: 1 / Chimera: Yes / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Streptococcus suis (bacteria)

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Macromolecule #1: KAP NTPase domain-containing protein

MacromoleculeName: KAP NTPase domain-containing protein / type: protein_or_peptide / ID: 1 / Number of copies: 7 / Enantiomer: LEVO
Source (natural)Organism: Streptococcus suis (bacteria)
Molecular weightTheoretical: 106.413305 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MTKINWEKYK KVIKKEFSEK EETEEVKNYI FSSQLDKVNL ILEHMDTVGG IHSRNIAITG DRGTGKTSFI ETLKLVLEKQ NYYVFDIVS PTVLSSHLNI LEIVISSIYR EIDQFIDSHD VHDRGRLIQH LKKVMNAIAV EKKQSDYFKQ SKPEIEMLTD L SHRTFLDE ...String:
MTKINWEKYK KVIKKEFSEK EETEEVKNYI FSSQLDKVNL ILEHMDTVGG IHSRNIAITG DRGTGKTSFI ETLKLVLEKQ NYYVFDIVS PTVLSSHLNI LEIVISSIYR EIDQFIDSHD VHDRGRLIQH LKKVMNAIAV EKKQSDYFKQ SKPEIEMLTD L SHRTFLDE EIKELFCYFK KVLNNRQDSC KEVIKDLVLI IDDLDLVENN LVYDLLRDIQ HYLDSQLIVI FAYKEGQLEQ SM FEHLAKG NEALLNHGVI DSNAIFGQIE RFLTKLVPLS NRIPLFKQDE LLNKTIGEFL ASLDPSYGVG ENLEFITKDS EKN KNNLTI REWFYESIFY RTNLKLDPID IREEASRLMP KTLREMVQLC EELHSMQVIT RSMDKLAGVE GLRKNIGAFR RYIG YKNST YFNLATMEFF QKWELAESHQ ANYLAYHFLM SYYQESFEQN QKLGYPLNLS KSGYPLTLRT MEPYNITLGD IYALM EELK YTEGISADTY YIVYILKVYY SLRLSELLYN VVLHHKLFVH VKEEATTFYM ATSTEELQIE KNHEQEATKL TDKEYR EHI MTAIEKVPAL QAYLELVNAQ FMPQNFNYDR SGSRDDDFYL ISWLKDDDLP EYSRLFKSLF LNSEVAAKGQ IQRNIGK RE SVFRYRNLYS YLPLQLTSAT FYKIDFLAFA IKADLLMYNV VRFVEEEGDT IPYFMSNMFH IDVFVRHNYN ENNNKGKF A YIAKQIVFGL WQGSNQRAHD LKHWYKSFDT VFGTKIEALH LLVDIAEQIK ISDKQTTDVS ALSEEQKRDE QAKKVAEKL AAIYHHIGMS RILSRLHQLP FIAEIKSNKE LLQHFSEAIV KLEKYASDTI NVGNLSQFRE SLKKIGQTYP SIQVLVDKLH RKQKLYVEF IQDFIETVNK LGEADESN

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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 TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.7000000000000001 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 42.02 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: OTHER
Final angle assignmentType: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.5 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 193305

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