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- PDB-4qqx: Crystal structure of T. fusca Cas3-ATP -

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Basic information

Entry
Database: PDB / ID: 4qqx
TitleCrystal structure of T. fusca Cas3-ATP
Components
  • CRISPR-associated helicase, Cas3 family
  • DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
KeywordsHydrolase/dna / CRISPR / Cas3 / hydrolase / helicase / hydrolase-DNA complex / helicase-DNA complex
Function / homology
Function and homology information


nuclease activity / defense response to virus / RNA helicase activity / ATP binding / metal ion binding / cytosol
Similarity search - Function
Hypothetical protein af1432 - #30 / Cas3, C-terminal / Cas3 C-terminal domain / Helicase Cas3, CRISPR-associated, core / Cas3, HD domain / CRISPR-associated Cas3-type HD domain / CRISPR-associated Cas3-type HD domain superfamily / HD Cas3-type domain profile. / Hypothetical protein af1432 / Helicase conserved C-terminal domain ...Hypothetical protein af1432 - #30 / Cas3, C-terminal / Cas3 C-terminal domain / Helicase Cas3, CRISPR-associated, core / Cas3, HD domain / CRISPR-associated Cas3-type HD domain / CRISPR-associated Cas3-type HD domain superfamily / HD Cas3-type domain profile. / Hypothetical protein af1432 / Helicase conserved C-terminal domain / helicase superfamily c-terminal domain / Helicase, C-terminal / P-loop containing nucleoside triphosphate hydrolase / Orthogonal Bundle / Mainly Alpha
Similarity search - Domain/homology
ADENOSINE-5'-TRIPHOSPHATE / : / DNA / DNA (> 10) / CRISPR-associated helicase, Cas3 family
Similarity search - Component
Biological speciesThermobifida fusca (bacteria)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.34 Å
AuthorsKe, A. / Huo, Y. / Nam, K.H.
CitationJournal: Nat.Struct.Mol.Biol. / Year: 2014
Title: Structures of CRISPR Cas3 offer mechanistic insights into Cascade-activated DNA unwinding and degradation.
Authors: Huo, Y. / Nam, K.H. / Ding, F. / Lee, H. / Wu, L. / Xiao, Y. / Farchione, M.D. / Zhou, S. / Rajashankar, K. / Kurinov, I. / Zhang, R. / Ke, A.
History
DepositionJun 30, 2014Deposition site: RCSB / Processing site: RCSB
Revision 1.0Aug 20, 2014Provider: repository / Type: Initial release
Revision 1.1Sep 24, 2014Group: Database references
Revision 1.2Sep 20, 2023Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / pdbx_struct_conn_angle / struct_conn / struct_ref_seq_dif / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_struct_conn_angle.ptnr1_auth_asym_id / _pdbx_struct_conn_angle.ptnr1_auth_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_asym_id / _pdbx_struct_conn_angle.ptnr1_label_atom_id / _pdbx_struct_conn_angle.ptnr1_label_comp_id / _pdbx_struct_conn_angle.ptnr1_label_seq_id / _pdbx_struct_conn_angle.ptnr2_auth_asym_id / _pdbx_struct_conn_angle.ptnr2_auth_seq_id / _pdbx_struct_conn_angle.ptnr2_label_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_asym_id / _pdbx_struct_conn_angle.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_asym_id / _pdbx_struct_conn_angle.ptnr3_label_atom_id / _pdbx_struct_conn_angle.ptnr3_label_comp_id / _pdbx_struct_conn_angle.ptnr3_label_seq_id / _pdbx_struct_conn_angle.value / _struct_conn.pdbx_dist_value / _struct_conn.ptnr1_auth_asym_id / _struct_conn.ptnr1_auth_comp_id / _struct_conn.ptnr1_auth_seq_id / _struct_conn.ptnr1_label_asym_id / _struct_conn.ptnr1_label_atom_id / _struct_conn.ptnr1_label_comp_id / _struct_conn.ptnr1_label_seq_id / _struct_conn.ptnr2_auth_asym_id / _struct_conn.ptnr2_auth_comp_id / _struct_conn.ptnr2_auth_seq_id / _struct_conn.ptnr2_label_asym_id / _struct_conn.ptnr2_label_atom_id / _struct_conn.ptnr2_label_comp_id / _struct_conn.ptnr2_label_seq_id / _struct_ref_seq_dif.details / _struct_site.pdbx_auth_asym_id / _struct_site.pdbx_auth_comp_id / _struct_site.pdbx_auth_seq_id

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: CRISPR-associated helicase, Cas3 family
B: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
C: CRISPR-associated helicase, Cas3 family
D: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
E: CRISPR-associated helicase, Cas3 family
F: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
G: CRISPR-associated helicase, Cas3 family
H: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)442,17220
Polymers439,6978
Non-polymers2,47512
Water00
1
C: CRISPR-associated helicase, Cas3 family
D: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)110,5435
Polymers109,9242
Non-polymers6193
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
2
A: CRISPR-associated helicase, Cas3 family
B: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)110,5435
Polymers109,9242
Non-polymers6193
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
3
E: CRISPR-associated helicase, Cas3 family
F: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)110,5435
Polymers109,9242
Non-polymers6193
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
4
G: CRISPR-associated helicase, Cas3 family
H: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)110,5435
Polymers109,9242
Non-polymers6193
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
5
A: CRISPR-associated helicase, Cas3 family
B: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
C: CRISPR-associated helicase, Cas3 family
D: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)221,08610
Polymers219,8484
Non-polymers1,2386
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area11630 Å2
ΔGint-98 kcal/mol
Surface area72360 Å2
MethodPISA
6
E: CRISPR-associated helicase, Cas3 family
F: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
G: CRISPR-associated helicase, Cas3 family
H: DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)221,08610
Polymers219,8484
Non-polymers1,2386
Water0
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area11240 Å2
ΔGint-95 kcal/mol
Surface area72240 Å2
MethodPISA
Unit cell
Length a, b, c (Å)87.250, 222.807, 125.087
Angle α, β, γ (deg.)90.000, 104.100, 90.000
Int Tables number4
Space group name H-MP1211

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Components

#1: Protein
CRISPR-associated helicase, Cas3 family


Mass: 106210.609 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Thermobifida fusca (bacteria) / Strain: YX / Gene: Tfu_1593 / Plasmid: pET28b / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3)star / References: UniProt: Q47PJ0
#2: DNA chain
DNA (5'-D(P*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*AP*A)-3')


Mass: 3713.524 Da / Num. of mol.: 4 / Source method: obtained synthetically
#3: Chemical
ChemComp-FE / FE (III) ION


Mass: 55.845 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Fe
#4: Chemical
ChemComp-ATP / ADENOSINE-5'-TRIPHOSPHATE


Mass: 507.181 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C10H16N5O13P3 / Comment: ATP, energy-carrying molecule*YM

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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: 2.68 Å3/Da / Density % sol: 54.14 %
Crystal growTemperature: 294 K / Method: vapor diffusion, hanging drop / pH: 6.5
Details: 100 mM MES, pH 6.5, and 5% 10% (w/v) PEG 8000, VAPOR DIFFUSION, HANGING DROP, temperature 294.0K

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Data collection

Diffraction
IDMean temperature (K)Crystal-ID
11001
21
Diffraction source
SourceSiteBeamlineID
SYNCHROTRONAPS 24-ID-C1
SYNCHROTRONCHESS A12
Detector
TypeIDDetector
ADSC QUANTUM 3151CCD
ADSC QUANTUM 2102CCD
RadiationMonochromator: Si 111 CHANNEL / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthRelative weight: 1
ReflectionResolution: 3.34→50 Å / Num. all: 66466 / Num. obs: 66466 / % possible obs: 99.21 % / Observed criterion σ(F): 2 / Observed criterion σ(I): 2 / Redundancy: 2.2 % / Biso Wilson estimate: 92.53 Å2 / Rmerge(I) obs: 0.064 / Net I/σ(I): 9.3
Reflection shellResolution: 3.34→3.49 Å / Redundancy: 2.2 % / Rmerge(I) obs: 0.412 / Mean I/σ(I) obs: 2.6 / % possible all: 99.6

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Processing

Software
NameVersionClassificationNB
PHENIX1.8.2_1309refinement
PDB_EXTRACT3.14data extraction
HKL-2000data collection
HKL-2000data reduction
HKL-2000data scaling
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 4QQW

4qqw


Resolution: 3.34→46.98 Å / FOM work R set: 0.8403 / SU ML: 0.37 / σ(F): 1.34 / Phase error: 23.53 / Stereochemistry target values: ML
RfactorNum. reflection% reflectionSelection details
Rfree0.2266 1618 2.43 %random
Rwork0.1719 ---
obs0.1733 66466 99.21 %-
all-66466 --
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 252.05 Å2 / Biso mean: 108.47 Å2 / Biso min: 20 Å2
Refinement stepCycle: LAST / Resolution: 3.34→46.98 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms27906 788 132 0 28826
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.01529663
X-RAY DIFFRACTIONf_angle_d1.56140672
X-RAY DIFFRACTIONf_chiral_restr0.0884556
X-RAY DIFFRACTIONf_plane_restr0.0065120
X-RAY DIFFRACTIONf_dihedral_angle_d18.78910861
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Total num. of bins used: 12

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
3.34-3.43830.3111340.244153835517100
3.4383-3.54920.29741360.234154215557100
3.5492-3.6760.26961340.208554185552100
3.676-3.82310.28311360.1954155551100
3.8231-3.99710.26511350.182754195554100
3.9971-4.20770.21831340.165154135547100
4.2077-4.47110.23461370.152554215558100
4.4711-4.8160.21941350.145554375572100
4.816-5.30010.19171340.15225380551499
5.3001-6.06560.25681350.17815416555199
6.0656-7.63670.21561340.18065397553199
7.6367-46.98510.1741340.15365328546296

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