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- PDB-5web: Crystal structure of the influenza virus PA endonuclease in compl... -

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Basic information

Entry
Database: PDB / ID: 5web
TitleCrystal structure of the influenza virus PA endonuclease in complex with inhibitor 10e (SRI-30024)
ComponentsPolymerase acidic protein
KeywordsHYDROLASE/HYDROLASE INHIBITOR / Virus / Nuclease / Transcription / Cap-snatching / HYDROLASE / HYDROLASE-HYDROLASE INHIBITOR complex
Function / homology
Function and homology information


cap snatching / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / endonuclease activity / Hydrolases; Acting on ester bonds / host cell cytoplasm / symbiont-mediated suppression of host gene expression / viral translational frameshifting / viral RNA genome replication / DNA-templated transcription / host cell nucleus ...cap snatching / symbiont-mediated suppression of host mRNA transcription via inhibition of RNA polymerase II activity / endonuclease activity / Hydrolases; Acting on ester bonds / host cell cytoplasm / symbiont-mediated suppression of host gene expression / viral translational frameshifting / viral RNA genome replication / DNA-templated transcription / host cell nucleus / RNA binding / metal ion binding
Similarity search - Function
Influenza RNA-dependent RNA polymerase subunit PA, endonuclease domain / Restriction Endonuclease / Polymerase acidic protein / Influenza RNA-dependent RNA polymerase subunit PA / Influenza RNA-dependent RNA polymerase subunit PA, endonuclease domain / Influenza RNA-dependent RNA polymerase subunit PA / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
Chem-KU5 / : / Polymerase acidic protein
Similarity search - Component
Biological speciesInfluenza A virus
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.254 Å
AuthorsKumar, G. / White, S.W.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI098757 United States
St. Jude Children's Research Hospital (ALSAC) United States
St. Jude Children's Research Hospital (ALSAC) United States
CitationJournal: Sci Rep / Year: 2017
Title: Protein-Structure Assisted Optimization of 4,5-Dihydroxypyrimidine-6-Carboxamide Inhibitors of Influenza Virus Endonuclease.
Authors: Beylkin, D. / Kumar, G. / Zhou, W. / Park, J. / Jeevan, T. / Lagisetti, C. / Harfoot, R. / Webby, R.J. / White, S.W. / Webb, T.R.
History
DepositionJul 8, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 3, 2018Provider: 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 / Derived calculations
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_struct_conn_angle / struct_conn
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_comp_id / _pdbx_struct_conn_angle.ptnr1_auth_seq_id / _pdbx_struct_conn_angle.ptnr1_label_alt_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.ptnr3_auth_comp_id / _pdbx_struct_conn_angle.ptnr3_auth_seq_id / _pdbx_struct_conn_angle.ptnr3_label_alt_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.value / _struct_conn.pdbx_dist_value / _struct_conn.pdbx_ptnr2_label_alt_id / _struct_conn.ptnr1_label_atom_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

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Polymerase acidic protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)24,0276
Polymers23,1481
Non-polymers8795
Water28816
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration, Protein elutes as a monomer
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area530 Å2
ΔGint-35 kcal/mol
Surface area9170 Å2
MethodPISA
Unit cell
Length a, b, c (Å)90.793, 90.793, 135.062
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number97
Space group name H-MI422

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Components

#1: Protein Polymerase acidic protein


Mass: 23148.344 Da / Num. of mol.: 1
Mutation: Loop 51-72 is replaced with a GGS linker, N-Terminal has His-tag residues
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Influenza A virus / Strain: swl A/California/04/2009 H1N1 / Gene: PA / Plasmid: pET28a / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: C3W5S0
#2: Chemical ChemComp-MN / MANGANESE (II) ION


Mass: 54.938 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mn
#3: Chemical ChemComp-KU5 / 2-[(2S)-1-{[(2-chlorophenyl)sulfanyl]acetyl}pyrrolidin-2-yl]-5-hydroxy-6-oxo-N-[2-(phenylsulfonyl)ethyl]-1,6-dihydropyrimidine-4-carboxamide / SRI-30024


Mass: 577.072 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C25H25ClN4O6S2 / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical ChemComp-SO4 / SULFATE ION


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


Mass: 18.015 Da / Num. of mol.: 16 / 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: 3.01 Å3/Da / Density % sol: 59.08 %
Crystal growTemperature: 291 K / Method: vapor diffusion, hanging drop / pH: 7.8
Details: 0.1 M HEPES pH 7.8, 1.0 M AmSO4, 10 mM MnCl2, 10 mM MgCl2, 1% PVP K15

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Data collection

DiffractionMean temperature: 100 K
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 22-BM / Wavelength: 0.97903 Å
DetectorType: MARMOSAIC 225 mm CCD / Detector: CCD / Date: Feb 9, 2017
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.97903 Å / Relative weight: 1
ReflectionResolution: 2.25→50 Å / Num. obs: 13682 / % possible obs: 99.8 % / Redundancy: 7 % / Biso Wilson estimate: 60.72 Å2 / Rmerge(I) obs: 0.059 / Rpim(I) all: 0.024 / Rrim(I) all: 0.064 / Χ2: 1.232 / Net I/σ(I): 11.3 / Num. measured all: 95657
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsCC1/2Rpim(I) allRrim(I) allΧ2% possible all
2.25-2.336.81.1080.7260.4491.1980.90399.8
2.33-2.427.10.7880.8780.3120.8480.941100
2.42-2.537.10.5210.9390.2060.5610.93199.9
2.53-2.677.10.3620.960.1430.390.96299.9
2.67-2.837.10.2550.9780.1010.2741.02799.9
2.83-3.057.10.1410.9930.0560.1521.10399.9
3.05-3.367.10.0880.9960.0350.0941.34599.9
3.36-3.8570.0630.9970.0250.0681.67100
3.85-4.856.90.0560.9970.0230.0611.82699.8
4.85-506.50.0340.9990.0140.0381.59399.2

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Processing

Software
NameVersionClassification
HKL-2000data scaling
PHENIXrefinement
PDB_EXTRACT3.22data extraction
HKL-2000data reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 2.254→38.885 Å / SU ML: 0.26 / Cross valid method: FREE R-VALUE / σ(F): 1.34 / Phase error: 31.95
RfactorNum. reflection% reflection
Rfree0.2546 734 5.37 %
Rwork0.2192 --
obs0.2212 13668 99.69 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso max: 154.62 Å2 / Biso mean: 90.0834 Å2 / Biso min: 54.72 Å2
Refinement stepCycle: final / Resolution: 2.254→38.885 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1409 0 88 16 1513
Biso mean--98.67 89.53 -
Num. residues----180
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0081529
X-RAY DIFFRACTIONf_angle_d0.9252073
X-RAY DIFFRACTIONf_chiral_restr0.064214
X-RAY DIFFRACTIONf_plane_restr0.005265
X-RAY DIFFRACTIONf_dihedral_angle_d4.331924
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 5

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
2.2543-2.42840.31341300.273125352665100
2.4284-2.67270.27541400.248525672707100
2.6727-3.05930.28391630.256725342697100
3.0593-3.85390.25671340.24226032737100
3.8539-38.89050.24211670.1952695286299
Refinement TLS params.Method: refined / Origin x: 202.4293 Å / Origin y: -310.5897 Å / Origin z: 693.2522 Å
111213212223313233
T0.5955 Å20.1435 Å20.1286 Å2-1.021 Å2-0.4465 Å2--0.7892 Å2
L2.0405 °20.7498 °20.5447 °2-5.0003 °2-1.887 °2--3.5734 °2
S-0.443 Å °-0.5574 Å °0.2689 Å °0.523 Å °0.0276 Å °0.667 Å °-0.1796 Å °-1.1028 Å °0.4003 Å °
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1allA-3 - 195
2X-RAY DIFFRACTION1allB1 - 2
3X-RAY DIFFRACTION1allC1 - 18
4X-RAY DIFFRACTION1allD1
5X-RAY DIFFRACTION1allD1
6X-RAY DIFFRACTION1allE1 - 2

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