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- PDB-8y05: Crystal structure of LbCas12a in complex with crRNA and 9nt target DNA -

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Basic information

Entry
Database: PDB / ID: 8y05
TitleCrystal structure of LbCas12a in complex with crRNA and 9nt target DNA
Components
  • DNA (5'-D(*CP*GP*TP*CP*CP*TP*TP*TP*AP*TP*T)-3')
  • DNA (5'-D(P*CP*TP*GP*GP*AP*TP*GP*CP*GP*TP*AP*AP*AP*GP*GP*AP*CP*G)-3')
  • LbCas12a
  • RNA (29-MER)
KeywordsRNA BINDING PROTEIN / CRISPR / Cas12a / crRNA / Complex
Function / homology
Function and homology information


: / CRISPR-associated endonuclease Cpf1 PI domain / : / CRISPR-associated endonuclease Cpf1 REC2 domain / CRISPR-associated endonuclease Cas12a / Cas12a, REC1 domain / Cas12a, RuvC nuclease domain / Cas12a, nuclease domain / Alpha helical recognition lobe domain / Nuclease domain / RuvC nuclease domain
Similarity search - Domain/homology
DNA / DNA (> 10) / RNA / RNA (> 10) / LbCas12a
Similarity search - Component
Biological speciesLachnospiraceae bacterium ND2006 (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.1 Å
AuthorsLin, X. / Chen, J. / Liu, L.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)32022047 China
CitationJournal: Nucleic Acids Res. / Year: 2025
Title: DNA target binding-induced pre-crRNA processing in type II and V CRISPR-Cas systems.
Authors: Chen, J. / Lin, X. / Xiang, W. / Chen, Y. / Zhao, Y. / Huang, L. / Liu, L.
History
DepositionJan 22, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 1, 2025Provider: repository / Type: Initial release
Revision 1.1Feb 19, 2025Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.year / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: LbCas12a
B: RNA (29-MER)
C: DNA (5'-D(P*CP*TP*GP*GP*AP*TP*GP*CP*GP*TP*AP*AP*AP*GP*GP*AP*CP*G)-3')
D: DNA (5'-D(*CP*GP*TP*CP*CP*TP*TP*TP*AP*TP*T)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)165,6135
Polymers165,5894
Non-polymers241
Water1086
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area12270 Å2
ΔGint-97 kcal/mol
Surface area59860 Å2
MethodPISA
Unit cell
Length a, b, c (Å)154.764, 154.764, 210.611
Angle α, β, γ (deg.)90.000, 90.000, 120.000
Int Tables number152
Space group name H-MP3121
Space group name HallP312"
Symmetry operation#1: x,y,z
#2: -y,x-y,z+1/3
#3: -x+y,-x,z+2/3
#4: x-y,-y,-z+2/3
#5: -x,-x+y,-z+1/3
#6: y,x,-z

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Components

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DNA chain , 2 types, 2 molecules CD

#3: DNA chain DNA (5'-D(P*CP*TP*GP*GP*AP*TP*GP*CP*GP*TP*AP*AP*AP*GP*GP*AP*CP*G)-3')


Mass: 5605.642 Da / Num. of mol.: 1 / Source method: obtained synthetically
Source: (synth.) Lachnospiraceae bacterium ND2006 (bacteria)
#4: DNA chain DNA (5'-D(*CP*GP*TP*CP*CP*TP*TP*TP*AP*TP*T)-3')


Mass: 3290.157 Da / Num. of mol.: 1 / Source method: obtained synthetically
Source: (synth.) Lachnospiraceae bacterium ND2006 (bacteria)

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Protein / RNA chain , 2 types, 2 molecules AB

#1: Protein LbCas12a


Mass: 143888.359 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lachnospiraceae bacterium ND2006 (bacteria)
Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A5S8WF58
#2: RNA chain RNA (29-MER)


Mass: 12804.658 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lachnospiraceae bacterium ND2006 (bacteria)
Production host: Escherichia coli BL21(DE3) (bacteria)

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Non-polymers , 2 types, 7 molecules

#5: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 6 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestN
Has protein modificationN

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Experimental details

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Experiment

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

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Sample preparation

CrystalDensity Matthews: 4.4 Å3/Da / Density % sol: 72.03 %
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop / Details: 0.1 M NaAc, pH 5.4, 8% PEG 2000MME

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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: Oct 22, 2022
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97915 Å / Relative weight: 1
ReflectionResolution: 3.1→50 Å / Num. obs: 53476 / % possible obs: 100 % / Redundancy: 9.1 % / Biso Wilson estimate: 62.52 Å2 / Rpim(I) all: 0.065 / Net I/σ(I): 11.25
Reflection shellResolution: 3.1→3.15 Å / Redundancy: 6.7 % / Mean I/σ(I) obs: 2 / Num. unique obs: 2636 / Rpim(I) all: 0.341

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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
Starting model: 5XUZ

5xuz


Resolution: 3.1→49.01 Å / SU ML: 0.3468 / Cross valid method: FREE R-VALUE / σ(F): 1.41 / Phase error: 22.9813
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.2345 2238 4.9 %
Rwork0.1972 43410 -
obs0.199 45648 85.37 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 69.52 Å2
Refinement stepCycle: LAST / Resolution: 3.1→49.01 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms9841 1203 1 6 11051
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.01111390
X-RAY DIFFRACTIONf_angle_d1.30215592
X-RAY DIFFRACTIONf_chiral_restr0.06691698
X-RAY DIFFRACTIONf_plane_restr0.00711789
X-RAY DIFFRACTIONf_dihedral_angle_d19.13261936
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.1-3.170.3828540.32081046X-RAY DIFFRACTION33.59
3.17-3.240.3044560.28541314X-RAY DIFFRACTION41.62
3.24-3.320.32881030.25011549X-RAY DIFFRACTION50.41
3.32-3.410.2821300.2431949X-RAY DIFFRACTION62.53
3.41-3.510.25311240.2362588X-RAY DIFFRACTION81.74
3.51-3.630.29971270.23943023X-RAY DIFFRACTION94.97
3.63-3.750.25141540.22733098X-RAY DIFFRACTION99.48
3.75-3.90.2731480.20273199X-RAY DIFFRACTION100
3.91-4.080.23061750.19043155X-RAY DIFFRACTION99.91
4.08-4.30.22131590.1793173X-RAY DIFFRACTION99.97
4.3-4.570.18691600.16633167X-RAY DIFFRACTION99.97
4.57-4.920.20721540.15863171X-RAY DIFFRACTION100
4.92-5.410.19951580.1693220X-RAY DIFFRACTION100
5.41-6.20.22371590.17943215X-RAY DIFFRACTION100
6.2-7.80.221850.19583226X-RAY DIFFRACTION99.97
7.8-49.010.22781920.19813317X-RAY DIFFRACTION98.9

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