[English] 日本語
Yorodumi
- PDB-7sba: Structure of type I-D Cascade bound to a dsDNA target -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 7sba
TitleStructure of type I-D Cascade bound to a dsDNA target
Components
  • Cas10d
  • Cas11d
  • Cas5d
  • Cas7d
  • DNA non-target strand
  • DNA target strand
  • crRNA
KeywordsDNA BINDING PROTEIN/DNA/RNA / CRISPR / Complex / Ribonucleoprotein complex / type I-D / type ID / type I / cyanobacteria / synechocystis / RNA binding protein / DNA binding protein / DNA BINDING PROTEIN-DNA-RNA complex
Function / homology
Function and homology information


CRISPR-associated protein Csc1 / CRISPR associated protein Csc3 / CRISPR-associated protein Csc2 / Csc2 Crispr
Similarity search - Domain/homology
: / DNA / DNA (> 10) / RNA / RNA (> 10) / Slr7013 protein / Slr7012 protein / CRISPR-associated protein Csc3
Similarity search - Component
Biological speciesSynechocystis sp. PCC 6803 (bacteria)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsSchwartz, E.A. / Taylor, D.W.
Funding support United States, 1items
OrganizationGrant numberCountry
Welch FoundationF-1938 United States
CitationJournal: Nat Commun / Year: 2022
Title: Structural rearrangements allow nucleic acid discrimination by type I-D Cascade.
Authors: Evan A Schwartz / Tess M McBride / Jack P K Bravo / Daniel Wrapp / Peter C Fineran / Robert D Fagerlund / David W Taylor /
Abstract: CRISPR-Cas systems are adaptive immune systems that protect prokaryotes from foreign nucleic acids, such as bacteriophages. Two of the most prevalent CRISPR-Cas systems include type I and type III. ...CRISPR-Cas systems are adaptive immune systems that protect prokaryotes from foreign nucleic acids, such as bacteriophages. Two of the most prevalent CRISPR-Cas systems include type I and type III. Interestingly, the type I-D interference proteins contain characteristic features of both type I and type III systems. Here, we present the structures of type I-D Cascade bound to both a double-stranded (ds)DNA and a single-stranded (ss)RNA target at 2.9 and 3.1 Å, respectively. We show that type I-D Cascade is capable of specifically binding ssRNA and reveal how PAM recognition of dsDNA targets initiates long-range structural rearrangements that likely primes Cas10d for Cas3' binding and subsequent non-target strand DNA cleavage. These structures allow us to model how binding of the anti-CRISPR protein AcrID1 likely blocks target dsDNA binding via competitive inhibition of the DNA substrate engagement with the Cas10d active site. This work elucidates the unique mechanisms used by type I-D Cascade for discrimination of single-stranded and double stranded targets. Thus, our data supports a model for the hybrid nature of this complex with features of type III and type I systems.
History
DepositionSep 24, 2021Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 8, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 5, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: Cas7d
B: Cas7d
C: Cas7d
D: Cas7d
E: Cas7d
F: Cas7d
G: Cas7d
H: Cas5d
I: Cas10d
J: Cas11d
K: Cas11d
X: DNA target strand
Y: DNA non-target strand
Z: crRNA


Theoretical massNumber of molelcules
Total (without water)453,28014
Polymers453,28014
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, Gel filtration followed by SDS-PAGE analysis and mass spectrometry. Cryo-electron microscopy was performed after verification of complex assembly and purification.
TypeNameSymmetry operationNumber
identity operation1_5551

-
Components

-
Protein , 4 types, 11 molecules ABCDEFGHIJK

#1: Protein
Cas7d


Mass: 36527.109 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechocystis sp. PCC 6803 (bacteria) / Gene: slr7012 / Production host: Escherichia coli (E. coli) / References: UniProt: Q6ZEI6
#2: Protein Cas5d


Mass: 28942.748 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechocystis sp. PCC 6803 (bacteria) / Gene: slr7013 / Production host: Escherichia coli (E. coli) / References: UniProt: Q6ZEI5
#3: Protein Cas10d


Mass: 112067.508 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechocystis sp. PCC 6803 (bacteria) / Gene: slr7011 / Production host: Escherichia coli (E. coli) / References: UniProt: Q6ZEI7
#4: Protein Cas11d


Mass: 17024.494 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechocystis sp. PCC 6803 (bacteria) / Gene: slr7011 / Production host: Escherichia coli (E. coli) / References: UniProt: Q6ZEI7

-
DNA chain , 2 types, 2 molecules XY

#5: DNA chain DNA target strand


Mass: 4884.222 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#6: DNA chain DNA non-target strand


Mass: 3947.581 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

-
RNA chain , 1 types, 1 molecules Z

#7: RNA chain crRNA


Mass: 13699.081 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Synechocystis sp. PCC 6803 (bacteria) / Production host: Escherichia coli (E. coli) / References: GenBank: CP073020

-
Experimental details

-
Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

-
Sample preparation

ComponentName: Type I-D Cascade bound to a dsDNA target / Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Molecular weightValue: 0.48 MDa / Experimental value: NO
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
1100 mMsodium chlorideNaCl1
220 mMHEPES1
31 mMDTT1
45 %Glycerol1
SpecimenConc.: 0.125 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: Monodisperse sample of elongated particles.
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K
Details: Blot force 0, blot time 4.5s, no wait time between sample application and blotting.

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 22500 X / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 3 sec. / Electron dose: 41.2 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1

-
Processing

EM software
IDNameVersionCategory
2Leginonimage acquisition
4WarpCTF correction
7Cootmodel fitting
9RELION3initial Euler assignment
12cryoSPARC23D reconstruction
13ISOLDEmodel refinement
Image processingDetails: Images were preprocessed and particles were picked via WARP, processing done using 2D classification in cryosparc v2, focused 3D classification in RELION, then Non-Uniform refinement in cryosparc v2.
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 4100000
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 336400 / Symmetry type: POINT
Atomic model buildingDetails: Model was built de novo using coot then flexibly refined using ISOLDE.

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbjlvh1.pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more