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- PDB-8y07: Crystal structure of LbCas12a in complex with crRNA and 13nt targ... -

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Basic information

Entry
Database: PDB / ID: 8y07
TitleCrystal structure of LbCas12a in complex with crRNA and 13nt target DNA
Components
  • DNA (5'-D(*CP*GP*TP*CP*CP*TP*TP*TP*AP*TP*T)-3')
  • DNA (5'-D(P*TP*TP*TP*AP*CP*TP*GP*GP*AP*TP*GP*CP*GP*TP*AP*AP*AP*GP*GP*AP*CP*G)-3')
  • LbCas12a
  • RNA (33-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: 2.85 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
E: LbCas12a
F: RNA (33-MER)
G: DNA (5'-D(P*TP*TP*TP*AP*CP*TP*GP*GP*AP*TP*GP*CP*GP*TP*AP*AP*AP*GP*GP*AP*CP*G)-3')
H: DNA (5'-D(*CP*GP*TP*CP*CP*TP*TP*TP*AP*TP*T)-3')
A: LbCas12a
B: RNA (33-MER)
C: DNA (5'-D(P*TP*TP*TP*AP*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)333,65712
Polymers333,6298
Non-polymers284
Water1629
1
E: LbCas12a
F: RNA (33-MER)
G: DNA (5'-D(P*TP*TP*TP*AP*CP*TP*GP*GP*AP*TP*GP*CP*GP*TP*AP*AP*AP*GP*GP*AP*CP*G)-3')
H: DNA (5'-D(*CP*GP*TP*CP*CP*TP*TP*TP*AP*TP*T)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)166,8286
Polymers166,8154
Non-polymers142
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area12730 Å2
ΔGint-101 kcal/mol
Surface area64130 Å2
MethodPISA
2
A: LbCas12a
B: RNA (33-MER)
C: DNA (5'-D(P*TP*TP*TP*AP*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)166,8286
Polymers166,8154
Non-polymers142
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area13230 Å2
ΔGint-109 kcal/mol
Surface area64330 Å2
MethodPISA
Unit cell
Length a, b, c (Å)121.664, 142.668, 202.687
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121
Space group name HallP2ac2ab
Symmetry operation#1: x,y,z
#2: x+1/2,-y+1/2,-z
#3: -x,y+1/2,-z+1/2
#4: -x+1/2,-y,z+1/2
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
111METMETARGARG(chain 'A' and (resid 1 through 91 or (resid 92...AE1 - 2841 - 284
121GLYGLYARGARG(chain 'A' and (resid 1 through 91 or (resid 92...AE291 - 1076291 - 1076
131PHEPHEVALVAL(chain 'A' and (resid 1 through 91 or (resid 92...AE1084 - 12261084 - 1226
211METMETARGARG(chain 'E' and (resid 1 through 129 or (resid 130...EA1 - 2841 - 284
221GLYGLYARGARG(chain 'E' and (resid 1 through 129 or (resid 130...EA291 - 1076291 - 1076
231PHEPHEVALVAL(chain 'E' and (resid 1 through 129 or (resid 130...EA1084 - 12261084 - 1226
112AAAAchain 'B'BF-20 - 121 - 33
212AAAAchain 'F'FB-20 - 121 - 33
113DTDTDGDGchain 'C'CG-12 - 91 - 22
213DTDTDGDGchain 'G'GC-12 - 91 - 22
114DCDCDTDTchain 'D'DH-9 - 11 - 11
214DCDCDTDTchain 'H'HD-9 - 11 - 11

NCS ensembles :
ID
1
2
3
4

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Components

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DNA chain , 2 types, 4 molecules GCHD

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


Mass: 6831.427 Da / Num. of mol.: 2 / 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.: 2 / Source method: obtained synthetically
Source: (synth.) Lachnospiraceae bacterium ND2006 (bacteria)

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Protein / RNA chain , 2 types, 4 molecules EAFB

#1: Protein LbCas12a


Mass: 143888.359 Da / Num. of mol.: 2
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 (33-MER)


Mass: 12804.658 Da / Num. of mol.: 2
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, 13 molecules

#5: Chemical
ChemComp-LI / LITHIUM ION


Mass: 6.941 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Li
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 9 / 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: 2.64 Å3/Da / Density % sol: 53.34 %
Crystal growTemperature: 289 K / Method: vapor diffusion, hanging drop
Details: 0.2 M Li2SO4, 30 mM Na2HPO4, 43.75 mM citric acid, 13% PEG 1000

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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: Mar 15, 2023
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97915 Å / Relative weight: 1
ReflectionResolution: 2.85→50 Å / Num. obs: 82713 / % possible obs: 100 % / Redundancy: 11.6 % / Biso Wilson estimate: 48.1 Å2 / Rpim(I) all: 0.072 / Net I/σ(I): 12
Reflection shellResolution: 2.85→2.9 Å / Redundancy: 10.7 % / Mean I/σ(I) obs: 3 / Num. unique obs: 4097 / Rpim(I) all: 0.27 / % possible all: 100

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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: 2.85→49.05 Å / SU ML: 0.3762 / Cross valid method: FREE R-VALUE / σ(F): 1.42 / Phase error: 26.3134
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
RfactorNum. reflection% reflection
Rfree0.2641 4043 4.97 %
Rwork0.2089 77275 -
obs0.2117 81318 98.26 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso mean: 54.9 Å2
Refinement stepCycle: LAST / Resolution: 2.85→49.05 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms19851 2740 4 9 22604
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.011523328
X-RAY DIFFRACTIONf_angle_d1.365232001
X-RAY DIFFRACTIONf_chiral_restr0.06713494
X-RAY DIFFRACTIONf_plane_restr0.00793628
X-RAY DIFFRACTIONf_dihedral_angle_d24.53329036
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.85-2.890.32811070.27072018X-RAY DIFFRACTION75.84
2.89-2.920.34151160.26442355X-RAY DIFFRACTION87.28
2.92-2.960.35651330.26822479X-RAY DIFFRACTION93.39
2.96-30.28991200.25862592X-RAY DIFFRACTION96.31
3-3.040.31791310.25122679X-RAY DIFFRACTION98.15
3.04-3.080.3261410.26452617X-RAY DIFFRACTION99.14
3.08-3.130.32931510.26472678X-RAY DIFFRACTION99.72
3.13-3.180.31360.24832685X-RAY DIFFRACTION99.82
3.18-3.230.3231520.23842663X-RAY DIFFRACTION100
3.23-3.280.29891440.23552702X-RAY DIFFRACTION99.93
3.28-3.340.30321360.22692664X-RAY DIFFRACTION100
3.34-3.410.28091560.222712X-RAY DIFFRACTION100
3.41-3.480.28651630.21942668X-RAY DIFFRACTION100
3.48-3.550.27131570.21762644X-RAY DIFFRACTION99.96
3.55-3.640.33941370.21232704X-RAY DIFFRACTION100
3.64-3.730.26251520.21182682X-RAY DIFFRACTION100
3.73-3.830.24131320.20042728X-RAY DIFFRACTION100
3.83-3.940.27621240.19322726X-RAY DIFFRACTION100
3.94-4.070.25251550.1882694X-RAY DIFFRACTION100
4.07-4.210.23041330.18392723X-RAY DIFFRACTION100
4.21-4.380.23911380.18282692X-RAY DIFFRACTION100
4.38-4.580.24791340.17612726X-RAY DIFFRACTION100
4.58-4.820.23151470.16552722X-RAY DIFFRACTION100
4.82-5.120.20351450.17032728X-RAY DIFFRACTION100
5.12-5.520.21251380.17882757X-RAY DIFFRACTION100
5.52-6.070.24791280.19052760X-RAY DIFFRACTION99.93
6.07-6.950.21661410.20192770X-RAY DIFFRACTION99.97
6.95-8.750.22751490.20392791X-RAY DIFFRACTION100
8.75-49.050.26041470.23372916X-RAY DIFFRACTION99.38

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