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Open data
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Basic information
Entry | Database: PDB / ID: 6xn7 | ||||||
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Title | Structure of the Lactococcus lactis Csm NTR CRISPR-Cas Complex | ||||||
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![]() | RNA BINDING PROTEIN/RNA / Type III-A CRISPR-Cas / Csm / RNA BINDING PROTEIN-RNA complex | ||||||
Function / homology | ![]() exonuclease activity / endonuclease activity / defense response to virus / RNA binding / ATP binding Similarity search - Function | ||||||
Biological species | ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.47 Å | ||||||
![]() | Rai, J. / Sridhara, S. / Li, H. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Structural and biochemical characterization of in vivo assembled Lactococcus lactis CRISPR-Csm complex. Authors: Sagar Sridhara / Jay Rai / Charlisa Whyms / Hemant Goswami / Huan He / Walter Woodside / Michael P Terns / Hong Li / ![]() ![]() Abstract: The small RNA-mediated immunity in bacteria depends on foreign RNA-activated and self RNA-inhibited enzymatic activities. The multi-subunit Type III-A CRISPR-Cas effector complex (Csm) exemplifies ...The small RNA-mediated immunity in bacteria depends on foreign RNA-activated and self RNA-inhibited enzymatic activities. The multi-subunit Type III-A CRISPR-Cas effector complex (Csm) exemplifies this principle and is in addition regulated by cellular metabolites such as divalent metals and ATP. Recognition of the foreign or cognate target RNA (CTR) triggers its single-stranded deoxyribonuclease (DNase) and cyclic oligoadenylate (cOA) synthesis activities. The same activities remain dormant in the presence of the self or non-cognate target RNA (NTR) that differs from CTR only in its 3'-protospacer flanking sequence (3'-PFS). Here we employ electron cryomicroscopy (cryoEM), functional assays, and comparative cross-linking to study in vivo assembled mesophilic Lactococcus lactis Csm (LlCsm) at the three functional states: apo, the CTR- and the NTR-bound. Unlike previously studied Csm complexes, we observed binding of 3'-PFS to Csm in absence of bound ATP and analyzed the structures of the four RNA cleavage sites. Interestingly, comparative crosslinking results indicate a tightening of the Csm3-Csm4 interface as a result of CTR but not NTR binding, reflecting a possible role of protein dynamics change during activation. | ||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 484.2 KB | Display | ![]() |
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PDB format | ![]() | 403.9 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Arichive directory | ![]() ![]() | HTTPS FTP |
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-Related structure data
Related structure data | ![]() 22269MC ![]() 6xn3C ![]() 6xn4C ![]() 6xn5C M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
-CRISPR-associated protein ... , 5 types, 10 molecules ABFHGIJCDE
#1: Protein | Mass: 87132.586 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: cas10 / Production host: ![]() ![]() | ||||
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#2: Protein | Mass: 33943.277 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: csm4 / Production host: ![]() ![]() | ||||
#3: Protein | Mass: 23825.160 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: csm3 / Production host: ![]() ![]() #4: Protein | | Mass: 40492.617 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: csm5 / Production host: ![]() ![]() #6: Protein | Mass: 15796.016 Da / Num. of mol.: 3 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Gene: csm2 / Production host: ![]() ![]() |
-RNA chain , 2 types, 2 molecules RT
#5: RNA chain | Mass: 11744.033 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Production host: ![]() ![]() |
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#7: RNA chain | Mass: 11938.096 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() Production host: ![]() ![]() |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Lactococcus lactis Csm CRISPR-Cas NTR Complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() ![]() |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: unspecified |
Vitrification | Cryogen name: ETHANE |
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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: ![]() |
Electron lens | Mode: OTHER / Cs: 2.7 mm / C2 aperture diameter: 70 µm |
Image recording | Electron dose: 60.14 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) |
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Processing
Software | Name: PHENIX / Version: 1.18.2_3874: / Classification: refinement | ||||||||||||||||||||||||
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.47 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 39220 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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