[English] 日本語
Yorodumi
- PDB-5u0a: CRISPR RNA-guided surveillance complex -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 5u0a
TitleCRISPR RNA-guided surveillance complex
Components
  • (CRISPR-associated protein, ...) x 4
  • Cse2Carbon diselenide
  • Nontarget Strand
  • Target Strand
  • crRNA
KeywordsIMMUNE SYSTEM / CRISPR-Cas / Cascacde / surveillance
Function / homology
Function and homology information


maintenance of CRISPR repeat elements / defense response to virus / RNA binding / identical protein binding
Similarity search - Function
CRISPR-associated protein, CT1975 / CT1975-like protein / CRISPR-associated protein, CasD / CRISPR-associated protein Cse2 / Cse2 superfamily / CRISPR-associated protein Cse2 (CRISPR_cse2) / CRISPR-associated protein Cse1 / CRISPR-associated protein Cse1 (CRISPR_cse1) / CRISPR-associated protein Cse3 / CRISPR associated protein ...CRISPR-associated protein, CT1975 / CT1975-like protein / CRISPR-associated protein, CasD / CRISPR-associated protein Cse2 / Cse2 superfamily / CRISPR-associated protein Cse2 (CRISPR_cse2) / CRISPR-associated protein Cse1 / CRISPR-associated protein Cse1 (CRISPR_cse1) / CRISPR-associated protein Cse3 / CRISPR associated protein / CRISPR_assoc / CRISPR-associated protein, Cas5 / CRISPR-associated protein (Cas_Cas5) / CRISPR-associated protein Cas5, N-terminal
Similarity search - Domain/homology
DNA / DNA (> 10) / RNA / RNA (> 10) / CRISPR-associated protein, Cse1 family / CRISPR-associated protein, Cse2 family / CRISPR-associated protein, Cse4 family / CRISPR-associated protein, Cas5e family / CRISPR-associated protein, Cse3 family
Similarity search - Component
Biological speciesThermobifida fusca (bacteria)
Thermobifida fusca YX (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsXiao, Y. / Luo, M. / Hayes, R.P. / Kim, J. / Ng, S. / Ding, F. / Liao, M. / Ke, A.
CitationJournal: Cell / Year: 2017
Title: Structure Basis for Directional R-loop Formation and Substrate Handover Mechanisms in Type I CRISPR-Cas System.
Authors: Yibei Xiao / Min Luo / Robert P Hayes / Jonathan Kim / Sherwin Ng / Fang Ding / Maofu Liao / Ailong Ke /
Abstract: Type I CRISPR systems feature a sequential dsDNA target searching and degradation process, by crRNA-displaying Cascade and nuclease-helicase fusion enzyme Cas3, respectively. Here we present two cryo- ...Type I CRISPR systems feature a sequential dsDNA target searching and degradation process, by crRNA-displaying Cascade and nuclease-helicase fusion enzyme Cas3, respectively. Here we present two cryo-EM snapshots of the Thermobifida fusca type I-E Cascade: (1) unwinding 11 bp of dsDNA at the seed-sequence region to scout for sequence complementarity, and (2) further unwinding of the entire protospacer to form a full R-loop. These structures provide the much-needed temporal and spatial resolution to resolve key mechanistic steps leading to Cas3 recruitment. In the early steps, PAM recognition causes severe DNA bending, leading to spontaneous DNA unwinding to form a seed-bubble. The full R-loop formation triggers conformational changes in Cascade, licensing Cas3 to bind. The same process also generates a bulge in the non-target DNA strand, enabling its handover to Cas3 for cleavage. The combination of both negative and positive checkpoints ensures stringent yet efficient target degradation in type I CRISPR-Cas systems.
History
DepositionNov 23, 2016Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 9, 2017Provider: repository / Type: Initial release
Revision 1.1Aug 16, 2017Group: Database references / Category: citation
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Jul 18, 2018Group: Data collection / Category: em_software / Item: _em_software.name
Revision 1.3Nov 6, 2019Group: Data collection / Other / Category: atom_sites / cell
Item: _atom_sites.fract_transf_matrix[1][1] / _atom_sites.fract_transf_matrix[2][2] ..._atom_sites.fract_transf_matrix[1][1] / _atom_sites.fract_transf_matrix[2][2] / _atom_sites.fract_transf_matrix[3][3] / _cell.Z_PDB / _cell.length_a / _cell.length_b / _cell.length_c
Revision 1.4Mar 13, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

-
Structure visualization

Movie
  • Deposited structure unit
  • Imaged by Jmol
  • Download
  • Superimposition on EM map
  • EMDB-8478
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: CRISPR-associated protein, Cse3 family
C: CRISPR-associated protein, Cse1 family
D: CRISPR-associated protein, Cse4 family
E: CRISPR-associated protein, Cse4 family
F: CRISPR-associated protein, Cse4 family
G: CRISPR-associated protein, Cse4 family
H: CRISPR-associated protein, Cse4 family
I: CRISPR-associated protein, Cse4 family
J: Cse2
K: crRNA
L: Cse2
M: Target Strand
N: CRISPR-associated protein, Cas5e family
O: Nontarget Strand


Theoretical massNumber of molelcules
Total (without water)464,31114
Polymers464,31114
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

-
Components

-
CRISPR-associated protein, ... , 4 types, 9 molecules ACDEFGHIN

#1: Protein CRISPR-associated protein, Cse3 family


Mass: 26327.938 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermobifida fusca (strain YX) (bacteria)
Strain: YX / Gene: Tfu_1588 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q47PJ5
#2: Protein CRISPR-associated protein, Cse1 family


Mass: 61433.297 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermobifida fusca (strain YX) (bacteria)
Strain: YX / Gene: Tfu_1592 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q47PJ1
#3: Protein
CRISPR-associated protein, Cse4 family / Cas7


Mass: 41043.043 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermobifida fusca (strain YX) (bacteria)
Strain: YX / Gene: Tfu_1590 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q47PJ3
#7: Protein CRISPR-associated protein, Cas5e family


Mass: 28279.260 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermobifida fusca (strain YX) (bacteria)
Strain: YX / Gene: Tfu_1589 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q47PJ4

-
DNA chain , 2 types, 2 molecules MO

#6: DNA chain Target Strand


Mass: 15251.741 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Thermobifida fusca YX (bacteria)
#8: DNA chain Nontarget Strand


Mass: 12076.723 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Thermobifida fusca YX (bacteria)

-
Protein / RNA chain , 2 types, 3 molecules JLK

#4: Protein Cse2 / Carbon diselenide


Mass: 27446.613 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermobifida fusca (strain YX) (bacteria)
Strain: YX / Gene: Tfu_1591 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q47PJ2
#5: RNA chain crRNA


Mass: 19790.793 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: RNA was prepared by in vitro transcription / Source: (synth.) Thermobifida fusca YX (bacteria)

-
Experimental details

-
Experiment

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

-
Sample preparation

ComponentName: CRISPR RNA-guided surveillance complex / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Thermobifida fusca (strain YX) (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Plasmid: pcdf
Buffer solutionpH: 7.5 / Details: 10 mM HEPES pH 7.5, 150 mM NaCl, 5 mM DTT
SpecimenConc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: 400 mesh Quantifoil holey carbon grid
VitrificationInstrument: GATAN CRYOPLUNGE 3 / Cryogen name: ETHANE / Humidity: 85 %

-
Electron microscopy imaging

Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
MicroscopyModel: FEI POLARA 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 31000 X / Calibrated defocus min: 1000 nm / Calibrated defocus max: 21000 nm / Cs: 2 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: GATAN LIQUID NITROGEN / Temperature (max): 105 K / Temperature (min): 80 K
Image recordingElectron dose: 8 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 1072

-
Processing

SoftwareName: PHENIX / Version: 1.11.1-2575_1692: / Classification: refinement
EM software
IDNameVersionCategory
1SPIDERparticle selection
2SAMUELparticle selection
3UcsfImager4image acquisition
5RELION1.4CTF correction
8Coot0.8.1model fitting
10SPIDERinitial Euler assignment
11RELION1.4final Euler assignment
12RELION1.4classification
13RELION1.43D reconstruction
14PHENIX1.11.1model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 303301
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 131292 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01330167
ELECTRON MICROSCOPYf_angle_d1.4241600
ELECTRON MICROSCOPYf_dihedral_angle_d18.35417710
ELECTRON MICROSCOPYf_chiral_restr0.0794700
ELECTRON MICROSCOPYf_plane_restr0.0084928

+
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://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