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Yorodumi- PDB-9c6c: cryoEM structure of CRISPR associated effector, CARF-Adenosine de... -
+Open data
-Basic information
Entry | Database: PDB / ID: 9c6c | ||||||
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Title | cryoEM structure of CRISPR associated effector, CARF-Adenosine deaminase 1, Cad1, in apo form with ATP (symmetric sites). | ||||||
Components | Adenosine deaminase domain-containing protein | ||||||
Keywords | ANTIVIRAL PROTEIN / Antiphage defense / CRISPR / Deamination / CARF-Adenosine deaminase / ATP | ||||||
Function / homology | Function and homology information inosine biosynthetic process / adenosine catabolic process / adenosine deaminase activity Similarity search - Function | ||||||
Biological species | Bacteroidales bacterium (bacteria) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å | ||||||
Authors | Majumder, P. / Patel, D.J. | ||||||
Funding support | United States, 1items
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Citation | Journal: Cell / Year: 2024 Title: The CRISPR-associated adenosine deaminase Cad1 converts ATP to ITP to provide antiviral immunity. Authors: Christian F Baca / Puja Majumder / James H Hickling / Linzhi Ye / Marianna Teplova / Sean F Brady / Dinshaw J Patel / Luciano A Marraffini / Abstract: Type III CRISPR systems provide immunity against genetic invaders through the production of cyclic oligo-adenylate (cA) molecules that activate effector proteins that contain CRISPR-associated ...Type III CRISPR systems provide immunity against genetic invaders through the production of cyclic oligo-adenylate (cA) molecules that activate effector proteins that contain CRISPR-associated Rossman fold (CARF) domains. Here, we characterized the function and structure of an effector in which the CARF domain is fused to an adenosine deaminase domain, CRISPR-associated adenosine deaminase 1 (Cad1). We show that upon binding of cA or cA to its CARF domain, Cad1 converts ATP to ITP, both in vivo and in vitro. Cryoelectron microscopy (cryo-EM) structural studies on full-length Cad1 reveal an hexameric assembly composed of a trimer of dimers, with bound ATP at inter-domain sites required for activity and ATP/ITP within deaminase active sites. Upon synthesis of cA during phage infection, Cad1 activation leads to a growth arrest of the host that prevents viral propagation. Our findings reveal that CRISPR-Cas systems employ a wide range of molecular mechanisms beyond nucleic acid degradation to provide adaptive immunity in prokaryotes. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 9c6c.cif.gz | 670.1 KB | Display | PDBx/mmCIF format |
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PDB format | pdb9c6c.ent.gz | 556.7 KB | Display | PDB format |
PDBx/mmJSON format | 9c6c.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 9c6c_validation.pdf.gz | 1.4 MB | Display | wwPDB validaton report |
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Full document | 9c6c_full_validation.pdf.gz | 1.5 MB | Display | |
Data in XML | 9c6c_validation.xml.gz | 108.8 KB | Display | |
Data in CIF | 9c6c_validation.cif.gz | 162 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/c6/9c6c ftp://data.pdbj.org/pub/pdb/validation_reports/c6/9c6c | HTTPS FTP |
-Related structure data
Related structure data | 45244MC 9c67C 9c68C 9c69C 9c6aC 9c6fC 9c77C 9cdbC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 67264.945 Da / Num. of mol.: 6 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Bacteroidales bacterium (bacteria) / Gene: DCM62_02910 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A3C0QUR5 #2: Chemical | ChemComp-MG / #3: Chemical | Has ligand of interest | Y | Has protein modification | N | |
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-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: apo Cad1 (CRISPR associated Adenosine deaminase 1) full length protein Type: COMPLEX / Details: It forms a trimer of dimers / Entity ID: #1 / Source: RECOMBINANT |
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Molecular weight | Value: 0.4 MDa / Experimental value: YES |
Source (natural) | Organism: Bacteriodale bacterium (bacteria) |
Source (recombinant) | Organism: Escherichia coli (E. coli) |
Buffer solution | pH: 8 Details: 25 mM Hepes pH 8, 200 mM NaCl, 2 mM Beta-mercaptoethanol and 5 % glycerol |
Specimen | Conc.: 1.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: Glow discharge 1 min, 15 mA wait time 100 % humidity, 12 s wait time, 2.5 s blot time and 0 blot force at 4 °C temperature Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-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: 2500 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Electron dose: 53 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K3 (6k x 4k) |
EM imaging optics | Energyfilter slit width: 20 eV |
-Processing
EM software | Name: PHENIX / Version: 1.20.1_4487: / Category: model refinement | ||||||||||||||||||||||||
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CTF correction | Type: NONE | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 106838 Details: The half maps were generated by the non-uniform refinemnet by cryoSparc. Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT / Space: REAL | ||||||||||||||||||||||||
Atomic model building | Source name: AlphaFold / Type: in silico model | ||||||||||||||||||||||||
Refine LS restraints |
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