[English] 日本語
Yorodumi
- PDB-8kal: Crystal structure of SpyCas9 in complex with sgRNA and 17nt target DNA -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: PDB / ID: 8kal
TitleCrystal structure of SpyCas9 in complex with sgRNA and 17nt target DNA
Components
  • CRISPR-associated endonuclease Cas9/Csn1
  • DNA (25-MER)
  • DNA (5'-D(*TP*TP*TP*AP*GP*GP*TP*AP*TP*TP*G)-3')
  • RNA (98-MER)
KeywordsRNA BINDING PROTEIN/RNA/DNA / Nuclease / RNA BINDING PROTEIN-RNA-DNA complex
Function / homology
Function and homology information


maintenance of CRISPR repeat elements / 3'-5' exonuclease activity / DNA endonuclease activity / defense response to virus / Hydrolases; Acting on ester bonds / DNA binding / RNA binding / metal ion binding
Similarity search - Function
CRISPR-associated endonuclease Cas9, bridge helix / Bridge helix of CRISPR-associated endonuclease Cas9 / CRISPR-associated endonuclease Cas9, PAM-interacting domain / CRISPR-associated endonuclease Cas9, REC lobe / REC lobe of CRISPR-associated endonuclease Cas9 / PAM-interacting domain of CRISPR-associated endonuclease Cas9 / : / Cas9 RuvC domain / HNH endonuclease / CRISPR-associated endonuclease Cas9 ...CRISPR-associated endonuclease Cas9, bridge helix / Bridge helix of CRISPR-associated endonuclease Cas9 / CRISPR-associated endonuclease Cas9, PAM-interacting domain / CRISPR-associated endonuclease Cas9, REC lobe / REC lobe of CRISPR-associated endonuclease Cas9 / PAM-interacting domain of CRISPR-associated endonuclease Cas9 / : / Cas9 RuvC domain / HNH endonuclease / CRISPR-associated endonuclease Cas9 / Cas9-type HNH domain / Cas9-type HNH domain profile. / HNH nuclease / Ribonuclease H superfamily
Similarity search - Domain/homology
DNA / DNA (> 10) / RNA / RNA (> 10) / CRISPR-associated endonuclease Cas9/Csn1
Similarity search - Component
Biological speciesStreptococcus pyogenes serotype M1 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.16 Å
AuthorsChen, Y. / Chen, J. / Liu, L.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32022047 China
CitationJournal: Nat.Biotechnol. / Year: 2025
Title: Trans-nuclease activity of Cas9 activated by DNA or RNA target binding.
Authors: Chen, J. / Chen, Y. / Huang, L. / Lin, X. / Chen, H. / Xiang, W. / Liu, L.
History
DepositionAug 3, 2023Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Jun 5, 2024Provider: repository / Type: Initial release
Revision 1.1Jun 12, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Apr 23, 2025Group: Database references / Structure summary / Category: citation / pdbx_entry_details
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.year / _pdbx_entry_details.has_protein_modification

-
Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

-
Assembly

Deposited unit
A: RNA (98-MER)
B: CRISPR-associated endonuclease Cas9/Csn1
C: DNA (25-MER)
D: DNA (5'-D(*TP*TP*TP*AP*GP*GP*TP*AP*TP*TP*G)-3')
E: RNA (98-MER)
G: CRISPR-associated endonuclease Cas9/Csn1
H: DNA (25-MER)
J: DNA (5'-D(*TP*TP*TP*AP*GP*GP*TP*AP*TP*TP*G)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)402,60210
Polymers402,4098
Non-polymers1922
Water1,13563
1
A: RNA (98-MER)
B: CRISPR-associated endonuclease Cas9/Csn1
C: DNA (25-MER)
D: DNA (5'-D(*TP*TP*TP*AP*GP*GP*TP*AP*TP*TP*G)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)201,3015
Polymers201,2054
Non-polymers961
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area18810 Å2
ΔGint-168 kcal/mol
Surface area80540 Å2
MethodPISA
2
E: RNA (98-MER)
G: CRISPR-associated endonuclease Cas9/Csn1
H: DNA (25-MER)
J: DNA (5'-D(*TP*TP*TP*AP*GP*GP*TP*AP*TP*TP*G)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)201,3015
Polymers201,2054
Non-polymers961
Water543
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area19000 Å2
ΔGint-155 kcal/mol
Surface area79440 Å2
MethodPISA
Unit cell
Length a, b, c (Å)362.049, 70.956, 200.100
Angle α, β, γ (deg.)90.000, 101.524, 90.000
Int Tables number5
Space group name H-MC121
Space group name HallC2y
Symmetry operation#1: x,y,z
#2: -x,y,-z
#3: x+1/2,y+1/2,z
#4: -x+1/2,y+1/2,-z
Noncrystallographic symmetry (NCS)NCS domain:
IDEns-ID
11
21
12
22
13
23
14
24

NCS domain segments:
Dom-IDComponent-IDEns-IDBeg auth comp-IDBeg label comp-IDEnd auth comp-IDEnd label comp-IDSelection detailsAuth asym-IDLabel asym-IDAuth seq-IDLabel seq-ID
111CCCCchain 'A'AA5 - 985 - 98
221CCCC(chain 'E' and resid 5 through 98)EE5 - 985 - 98
112LYSLYSARGARG(chain 'B' and (resid 4 through 1241 or (resid 1242...BB4 - 7654 - 765
122ASNASNASPASP(chain 'B' and (resid 4 through 1241 or (resid 1242...BB776 - 1012776 - 1012
132GLYGLYILEILE(chain 'B' and (resid 4 through 1241 or (resid 1242...BB1030 - 10501030 - 1050
142LYSLYSGLNGLN(chain 'B' and (resid 4 through 1241 or (resid 1242...BB1059 - 13641059 - 1364
252LYSLYSARGARG(chain 'G' and (resid 4 through 64 or (resid 65...GF4 - 7654 - 765
262ASNASNASPASP(chain 'G' and (resid 4 through 64 or (resid 65...GF776 - 1012776 - 1012
272GLYGLYILEILE(chain 'G' and (resid 4 through 64 or (resid 65...GF1030 - 10501030 - 1050
282LYSLYSGLNGLN(chain 'G' and (resid 4 through 64 or (resid 65...GF1059 - 13641059 - 1364
113DCDCDGDGchain 'C'CC1 - 251 - 25
223DCDCDGDGchain 'H'HG1 - 251 - 25
114DTDTDGDGchain 'D'DD2 - 121 - 11
224DTDTDGDGchain 'J'JH2 - 121 - 11

NCS ensembles :
ID
1
2
3
4

-
Components

-
DNA chain , 2 types, 4 molecules CHDJ

#3: DNA chain DNA (25-MER)


Mass: 7610.952 Da / Num. of mol.: 2 / Source method: obtained synthetically / Details: TS
Source: (synth.) Streptococcus pyogenes serotype M1 (bacteria)
#4: DNA chain DNA (5'-D(*TP*TP*TP*AP*GP*GP*TP*AP*TP*TP*G)-3')


Mass: 3394.230 Da / Num. of mol.: 2 / Source method: obtained synthetically / Details: NTS
Source: (synth.) Streptococcus pyogenes serotype M1 (bacteria)

-
RNA chain / Protein , 2 types, 4 molecules AEBG

#1: RNA chain RNA (98-MER)


Mass: 31610.740 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptococcus pyogenes serotype M1 (bacteria)
Production host: Escherichia coli DH5[alpha] (bacteria)
#2: Protein CRISPR-associated endonuclease Cas9/Csn1 / SpCas9 / SpyCas9


Mass: 158588.781 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptococcus pyogenes serotype M1 (bacteria)
Gene: cas9, csn1, SPy_1046 / Production host: Escherichia coli BL21(DE3) (bacteria)
References: UniProt: Q99ZW2, Hydrolases; Acting on ester bonds

-
Non-polymers , 2 types, 65 molecules

#5: Chemical ChemComp-SO4 / SULFATE ION


Mass: 96.063 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: SO4
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 63 / Source method: isolated from a natural source / Formula: H2O

-
Details

Has ligand of interestN
Has protein modificationN

-
Experimental details

-
Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

-
Sample preparation

CrystalDensity Matthews: 3.22 Å3/Da / Density % sol: 61.86 %
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop
Details: 0.2 M Ammonium sulfate, 0.1 M MES pH 6.0, 12.625% PEG8000

-
Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRF / Beamline: BL18U1 / Wavelength: 0.97915 Å
DetectorType: DECTRIS PILATUS 6M / Detector: PIXEL / Date: Mar 15, 2023
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97915 Å / Relative weight: 1
ReflectionResolution: 3.15→50.17 Å / Num. obs: 83075 / % possible obs: 99.1 % / Redundancy: 4.5 % / Biso Wilson estimate: 58.11 Å2 / Rpim(I) all: 0.187 / Net I/σ(I): 3
Reflection shellResolution: 3.15→3.2 Å / Num. unique obs: 4098 / Rpim(I) all: 0.54 / % possible all: 98

-
Processing

Software
NameVersionClassification
PHENIX1.17.1_3660refinement
HKL-30007.21data scaling
HKL-30007.21data reduction
Coot0.7.2.1model building
PHENIX1.17.1_3660phasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 3.16→50.17 Å / SU ML: 0.4583 / Cross valid method: FREE R-VALUE / σ(F): 1.99 / Phase error: 31.6112
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.2884 2796 4.94 %
Rwork0.2508 53831 -
obs0.2526 56627 65.56 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 59.6 Å2
Refinement stepCycle: LAST / Resolution: 3.16→50.17 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms21643 5498 10 63 27214
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.011728188
X-RAY DIFFRACTIONf_angle_d1.556739167
X-RAY DIFFRACTIONf_chiral_restr0.17724487
X-RAY DIFFRACTIONf_plane_restr0.00614062
X-RAY DIFFRACTIONf_dihedral_angle_d22.993911469
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.16-3.210.484270.2639205X-RAY DIFFRACTION4.85
3.22-3.270.3263130.2929311X-RAY DIFFRACTION7.75
3.27-3.340.3373240.3074422X-RAY DIFFRACTION10.44
3.34-3.40.3195350.3235518X-RAY DIFFRACTION12.98
3.4-3.480.3239490.3205773X-RAY DIFFRACTION19.27
3.48-3.560.3118700.32291125X-RAY DIFFRACTION27.67
3.56-3.650.3173770.30671566X-RAY DIFFRACTION38.96
3.65-3.750.29971200.28472359X-RAY DIFFRACTION57.15
3.75-3.860.27051510.26422866X-RAY DIFFRACTION71.22
3.86-3.980.30891640.2693348X-RAY DIFFRACTION81.71
3.98-4.120.34522010.26713735X-RAY DIFFRACTION91.51
4.12-4.290.33932370.25263848X-RAY DIFFRACTION95.07
4.29-4.480.27092150.24913952X-RAY DIFFRACTION96.57
4.48-4.720.29752170.24124058X-RAY DIFFRACTION98.41
4.72-5.020.28921940.23474044X-RAY DIFFRACTION98.95
5.02-5.40.27472040.23694097X-RAY DIFFRACTION98.99
5.4-5.950.26581870.24924118X-RAY DIFFRACTION99.1
5.95-6.80.30761960.27014125X-RAY DIFFRACTION99.38
6.8-8.570.27882230.23524135X-RAY DIFFRACTION99.27
8.57-50.170.2352120.21724226X-RAY DIFFRACTION97.35

+
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