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- PDB-6anw: Crystal structure of anti-CRISPR protein AcrF10 -

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Basic information

Entry
Database: PDB / ID: 6anw
TitleCrystal structure of anti-CRISPR protein AcrF10
Componentsanti-CRISPR protein AcrF10
KeywordsIMMUNE SYSTEM / Type I-F CRISPR-Cas system: Csy Cascade: Structure: anti-CRISPR protein: Inhibition of Csy complex: Genome editing tool
Function / homologyAcyl carrier protein
Function and homology information
Biological speciesShewanella xiamenensis (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / SAD / Resolution: 2.486 Å
AuthorsYang, H. / Patel, D.J.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute on Drug Abuse (NIH/NIDA)GM104962 United States
Memorial Sloan-Kettering Cancer Center Core GrantP30CA008748 United States
CitationJournal: Cell / Year: 2017
Title: Cryo-EM Structures Reveal Mechanism and Inhibition of DNA Targeting by a CRISPR-Cas Surveillance Complex.
Authors: Tai Wei Guo / Alberto Bartesaghi / Hui Yang / Veronica Falconieri / Prashant Rao / Alan Merk / Edward T Eng / Ashleigh M Raczkowski / Tara Fox / Lesley A Earl / Dinshaw J Patel / Sriram Subramaniam /
Abstract: Prokaryotic cells possess CRISPR-mediated adaptive immune systems that protect them from foreign genetic elements, such as invading viruses. A central element of this immune system is an RNA-guided ...Prokaryotic cells possess CRISPR-mediated adaptive immune systems that protect them from foreign genetic elements, such as invading viruses. A central element of this immune system is an RNA-guided surveillance complex capable of targeting non-self DNA or RNA for degradation in a sequence- and site-specific manner analogous to RNA interference. Although the complexes display considerable diversity in their composition and architecture, many basic mechanisms underlying target recognition and cleavage are highly conserved. Using cryoelectron microscopy (cryo-EM), we show that the binding of target double-stranded DNA (dsDNA) to a type I-F CRISPR system yersinia (Csy) surveillance complex leads to large quaternary and tertiary structural changes in the complex that are likely necessary in the pathway leading to target dsDNA degradation by a trans-acting helicase-nuclease. Comparison of the structure of the surveillance complex before and after dsDNA binding, or in complex with three virally encoded anti-CRISPR suppressors that inhibit dsDNA binding, reveals mechanistic details underlying target recognition and inhibition.
History
DepositionAug 14, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 25, 2017Provider: repository / Type: Initial release
Revision 1.1Dec 11, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.2Mar 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

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: anti-CRISPR protein AcrF10
B: anti-CRISPR protein AcrF10
C: anti-CRISPR protein AcrF10


Theoretical massNumber of molelcules
Total (without water)34,1393
Polymers34,1393
Non-polymers00
Water23413
1
A: anti-CRISPR protein AcrF10


Theoretical massNumber of molelcules
Total (without water)11,3801
Polymers11,3801
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
2
B: anti-CRISPR protein AcrF10


Theoretical massNumber of molelcules
Total (without water)11,3801
Polymers11,3801
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
3
C: anti-CRISPR protein AcrF10


Theoretical massNumber of molelcules
Total (without water)11,3801
Polymers11,3801
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
Unit cell
Length a, b, c (Å)73.774, 106.618, 95.349
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number20
Space group name H-MC2221

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Components

#1: Protein anti-CRISPR protein AcrF10


Mass: 11379.665 Da / Num. of mol.: 3 / Fragment: anti-CRISPR AcrF10
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Shewanella xiamenensis (bacteria) / Gene: SXM_1276 / Plasmid: pRSFDuet-SUMO / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: A0A073KP86
#2: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 13 / Source method: isolated from a natural source / Formula: H2O

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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: 2.78 Å3/Da / Density % sol: 55.8 %
Crystal growTemperature: 293 K / Method: evaporation / pH: 7.5 / Details: 0.1 M HEPES (pH 7.5) and 1.4 M tri-sodium citrate

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 24-ID-C / Wavelength: 0.9792 Å
DetectorType: ADSC QUANTUM 315 / Detector: CCD / Date: Oct 25, 2016 / Details: LR-Design detector positioner
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.9792 Å / Relative weight: 1
ReflectionResolution: 2.5→50 Å / Num. obs: 13323 / % possible obs: 98.9 % / Redundancy: 4.3 % / Biso Wilson estimate: 52.62 Å2 / Rmerge(I) obs: 0.113 / Rpim(I) all: 0.06 / Rrim(I) all: 0.128 / Χ2: 1.779 / Net I/σ(I): 7.6 / Num. measured all: 57879
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. unique obsCC1/2Rpim(I) allRrim(I) allΧ2% possible all
2.5-2.594.20.45812990.8550.2460.5230.55899.2
2.59-2.694.30.36913260.8970.1940.4190.63399
2.69-2.8240.26613110.930.1460.3050.71898.5
2.82-2.964.50.21513110.950.1110.2431.04399.7
2.96-3.154.60.16813120.9630.0870.191.35199.5
3.15-3.394.50.13613440.9780.070.1541.98998.8
3.39-3.734.10.1213190.9690.0670.1382.76399.2
3.73-4.274.60.10613440.9850.0540.122.91899.3
4.27-5.384.30.09213500.9880.0480.1052.57298
5.38-504.20.09614070.990.0520.113.04297.9

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Processing

Software
NameVersionClassification
PHENIX(1.11.1_2575: ???)refinement
HKL-2000data scaling
PDB_EXTRACT3.22data extraction
AutoSolphasing
RefinementMethod to determine structure: SAD / Resolution: 2.486→47.675 Å / SU ML: 0.4 / Cross valid method: FREE R-VALUE / σ(F): 1.38 / Phase error: 31.29 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2691 671 5.04 %
Rwork0.2176 --
obs0.2202 13306 98.12 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 2.486→47.675 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2292 0 0 13 2305
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0052325
X-RAY DIFFRACTIONf_angle_d0.6423146
X-RAY DIFFRACTIONf_dihedral_angle_d31.7856
X-RAY DIFFRACTIONf_chiral_restr0.055366
X-RAY DIFFRACTIONf_plane_restr0.003410
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.4864-2.67830.39371320.30422403X-RAY DIFFRACTION95
2.6783-2.94780.35591270.29392529X-RAY DIFFRACTION99
2.9478-3.37430.32061230.25312525X-RAY DIFFRACTION99
3.3743-4.25090.24731500.20582547X-RAY DIFFRACTION99
4.2509-47.68320.22811390.18182631X-RAY DIFFRACTION98
Refinement TLS params.Method: refined / Origin x: -12.1266 Å / Origin y: 17.5287 Å / Origin z: -10.556 Å
111213212223313233
T0.4218 Å2-0.0788 Å20.0343 Å2-0.4166 Å20.0723 Å2--0.3446 Å2
L1.8254 °21.5336 °20.1914 °2-2.0373 °20.25 °2---0.1403 °2
S-0.0787 Å °0.0474 Å °-0.187 Å °-0.2523 Å °0.0771 Å °-0.2298 Å °-0.0202 Å °-0.0729 Å °-0.002 Å °
Refinement TLS groupSelection details: all

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