+Open data
-Basic information
Entry | Database: PDB / ID: 8qlo | ||||||
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Title | CryoEM structure of the apo SPARTA (BabAgo/TIR-APAZ) complex | ||||||
Components |
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Keywords | IMMUNE SYSTEM / Prokaryotic Argonaute / TIR domain / RNA binding protein / DNA binding protein | ||||||
Biological species | Bacillales bacterium (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.57 Å | ||||||
Authors | Finocchio, G. / Koopal, B. / Potocnik, A. / Heijstek, C. / Jinek, M. / Swarts, D. | ||||||
Funding support | European Union, 1items
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Citation | Journal: Nucleic Acids Res / Year: 2024 Title: Target DNA-dependent activation mechanism of the prokaryotic immune system SPARTA. Authors: Giada Finocchio / Balwina Koopal / Ana Potocnik / Clint Heijstek / Adrie H Westphal / Martin Jinek / Daan C Swarts / Abstract: In both prokaryotic and eukaryotic innate immune systems, TIR domains function as NADases that degrade the key metabolite NAD+ or generate signaling molecules. Catalytic activation of TIR domains ...In both prokaryotic and eukaryotic innate immune systems, TIR domains function as NADases that degrade the key metabolite NAD+ or generate signaling molecules. Catalytic activation of TIR domains requires oligomerization, but how this is achieved varies in distinct immune systems. In the Short prokaryotic Argonaute (pAgo)/TIR-APAZ (SPARTA) immune system, TIR NADase activity is triggered upon guide RNA-mediated recognition of invading DNA by an unknown mechanism. Here, we describe cryo-EM structures of SPARTA in the inactive monomeric and target DNA-activated tetrameric states. The monomeric SPARTA structure reveals that in the absence of target DNA, a C-terminal tail of TIR-APAZ occupies the nucleic acid binding cleft formed by the pAgo and TIR-APAZ subunits, inhibiting SPARTA activation. In the active tetrameric SPARTA complex, guide RNA-mediated target DNA binding displaces the C-terminal tail and induces conformational changes in pAgo that facilitate SPARTA-SPARTA dimerization. Concurrent release and rotation of one TIR domain allow it to form a composite NADase catalytic site with the other TIR domain within the dimer, and generate a self-complementary interface that mediates cooperative tetramerization. Combined, this study provides critical insights into the structural architecture of SPARTA and the molecular mechanism underlying target DNA-dependent oligomerization and catalytic activation. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8qlo.cif.gz | 190.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8qlo.ent.gz | 148.7 KB | Display | PDB format |
PDBx/mmJSON format | 8qlo.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8qlo_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 8qlo_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 8qlo_validation.xml.gz | 39.8 KB | Display | |
Data in CIF | 8qlo_validation.cif.gz | 56.7 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ql/8qlo ftp://data.pdbj.org/pub/pdb/validation_reports/ql/8qlo | HTTPS FTP |
-Related structure data
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 57911.199 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacillales bacterium (bacteria) / Production host: Escherichia coli (E. coli) |
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#2: Protein | Mass: 53052.355 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacillales bacterium (bacteria) / Production host: Escherichia coli (E. coli) |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Complex of BabAgo with BabTIR-APAZ / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Source (natural) | Organism: Bacillales bacterium (bacteria) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 1000 nm |
Image recording | Electron dose: 58.005 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
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3D reconstruction | Resolution: 2.57 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 904721 / Symmetry type: POINT | ||||||||||||||||||||||||
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