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- PDB-4pmb: Crystal structure of Staphylcoccal nuclease variant Delta+PHS I92... -

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Basic information

Entry
Database: PDB / ID: 4pmb
TitleCrystal structure of Staphylcoccal nuclease variant Delta+PHS I92S at cryogenic temperature
ComponentsThermonuclease
KeywordsHYDROLASE / nuclease / hyperstable / pdTp / polar group
Function / homology
Function and homology information


micrococcal nuclease / endonuclease activity, active with either ribo- or deoxyribonucleic acids and producing 3'-phosphomonoesters / nucleic acid binding / extracellular region / membrane / metal ion binding
Similarity search - Function
OB fold (Dihydrolipoamide Acetyltransferase, E2P) - #90 / Thermonuclease family signature 1. / Thermonuclease active site / Thermonuclease family signature 2. / Staphylococcal nuclease (SNase-like), OB-fold / Staphylococcal nuclease homologue / Thermonuclease domain profile. / Staphylococcal nuclease homologues / SNase-like, OB-fold superfamily / OB fold (Dihydrolipoamide Acetyltransferase, E2P) ...OB fold (Dihydrolipoamide Acetyltransferase, E2P) - #90 / Thermonuclease family signature 1. / Thermonuclease active site / Thermonuclease family signature 2. / Staphylococcal nuclease (SNase-like), OB-fold / Staphylococcal nuclease homologue / Thermonuclease domain profile. / Staphylococcal nuclease homologues / SNase-like, OB-fold superfamily / OB fold (Dihydrolipoamide Acetyltransferase, E2P) / Beta Barrel / Mainly Beta
Similarity search - Domain/homology
THYMIDINE-3',5'-DIPHOSPHATE / Thermonuclease
Similarity search - Component
Biological speciesStaphylococcus aureus (bacteria)
MethodX-RAY DIFFRACTION / MOLECULAR REPLACEMENT / molecular replacement / Resolution: 1.8 Å
AuthorsSorenson, J.L. / Schlessman, J.L. / Garcia-Moreno, E.B.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)NIH-GM061597 United States
CitationJournal: to be published
Title: Crystal structure of Staphylcoccal nuclease variant Delta+PHS I92S at cryogenic temperature
Authors: Sorenson, J.L. / Schlessman, J.L. / Garcia-Moreno, E.B.
History
DepositionMay 20, 2014Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 11, 2014Provider: repository / Type: Initial release
Revision 1.1Feb 4, 2015Group: Derived calculations
Revision 1.2Feb 25, 2015Group: Derived calculations
Revision 1.3Sep 6, 2017Group: Author supporting evidence / Database references ...Author supporting evidence / Database references / Derived calculations / Other / Source and taxonomy
Category: citation / entity_src_gen ...citation / entity_src_gen / pdbx_audit_support / pdbx_database_status / pdbx_struct_assembly / pdbx_struct_conn_angle / pdbx_struct_oper_list
Item: _citation.journal_id_CSD / _entity_src_gen.pdbx_alt_source_flag ..._citation.journal_id_CSD / _entity_src_gen.pdbx_alt_source_flag / _pdbx_audit_support.funding_organization / _pdbx_database_status.pdb_format_compatible / _pdbx_struct_assembly.oligomeric_details / _pdbx_struct_oper_list.symmetry_operation
Revision 1.4Nov 22, 2017Group: Refinement description / Category: software / Item: _software.classification
Revision 1.5Dec 25, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.6Sep 27, 2023Group: Data collection / Database references / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_initial_refinement_model / refine_hist
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _refine_hist.number_atoms_total / _refine_hist.pdbx_number_atoms_nucleic_acid / _refine_hist.pdbx_number_atoms_protein

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Thermonuclease
hetero molecules


Theoretical massNumber of molelcules
Total (without water)16,5603
Polymers16,1171
Non-polymers4422
Water1,20767
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
Unit cell
Length a, b, c (Å)31.031, 59.964, 38.017
Angle α, β, γ (deg.)90.000, 93.630, 90.000
Int Tables number4
Space group name H-MP1211

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Components

#1: Protein Thermonuclease / TNase / Micrococcal nuclease / Staphylococcal nuclease


Mass: 16117.383 Da / Num. of mol.: 1 / Fragment: UNP residues 83-231
Mutation: Delta 44-49, G50F, V51N, I92S, P117G, H124L, S128A
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Staphylococcus aureus (bacteria) / Gene: nuc / Plasmid: pET24a+ / Production host: Escherichia coli (E. coli) / Strain (production host): BL21(DE3) / References: UniProt: P00644, micrococcal nuclease
#2: Chemical ChemComp-THP / THYMIDINE-3',5'-DIPHOSPHATE


Type: DNA linking / Mass: 402.188 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H16N2O11P2
#3: Chemical ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Ca
#4: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 67 / 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: 2.19 Å3/Da / Density % sol: 43.84 %
Crystal growTemperature: 277 K / Method: vapor diffusion, hanging drop / pH: 8
Details: 25% MPD, 25mM potassium phosphate, calcium chloride, pdTp

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Data collection

DiffractionMean temperature: 110 K
Diffraction sourceSource: SEALED TUBE / Type: OTHER / Wavelength: 1.54 Å
DetectorType: APEX II CCD / Detector: CCD / Date: May 7, 2013 / Details: mirrors
RadiationMonochromator: Si(111) / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.54 Å / Relative weight: 1
ReflectionResolution: 1.8→59.96 Å / Num. all: 12978 / Num. obs: 12978 / % possible obs: 99.9 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 9.08 % / Biso Wilson estimate: 27.2 Å2 / Rmerge(I) obs: 0.0347 / Net I/σ(I): 18.95 / Num. measured all: 120929
Reflection shell

Diffraction-ID: 1 / Rejects: _

Resolution (Å)Redundancy (%)Rmerge(I) obsNum. possibleNum. unique obsNet I/σ(I) obs% possible all
1.8-1.835.680.22664644643.54100
1.83-1.876.10.352749747499.7
1.87-1.916.280.2827267224.8399.4
1.91-1.966.50.2338248225.6599.8
1.96-2.016.840.2017457456.36100
2.01-2.066.930.1746666658.3599.8
2.06-2.127.050.1737287269.0699.7
2.12-2.197.460.14374774710.96100
2.19-2.267.750.13166166012.3999.8
2.26-2.348.220.11965465313.7999.8
2.34-2.448.550.10670670615.34100
2.44-2.559.190.09466166117.55100
2.55-2.699.690.08867767621.2299.9
2.69-2.8610.50.07966366323.26100
2.86-3.0911.440.0768368330.28100
3.09-3.4113.090.06167267236.58100
3.41-3.9213.830.05466566543.65100
3.92-4.9814.250.05165165149.39100
4.98-59.9614.280.04965065049.69100

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Phasing

PhasingMethod: molecular replacement
Phasing MRModel details: Phaser MODE: MR_AUTO
Highest resolutionLowest resolution
Rotation1.8 Å37.94 Å
Translation1.8 Å37.94 Å

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Processing

Software
NameVersionClassification
SAINT2008/2 for Windowsdata reduction
SAINTdata scaling
PHASERphasing
REFMAC5.7.0029refinement
PDB_EXTRACT3.14data extraction
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: PDB entry 3BDC

3bdc


Resolution: 1.8→37.94 Å / Cor.coef. Fo:Fc: 0.954 / Cor.coef. Fo:Fc free: 0.938 / WRfactor Rfree: 0.2116 / WRfactor Rwork: 0.1872 / FOM work R set: 0.8198 / SU B: 5.237 / SU ML: 0.089 / SU R Cruickshank DPI: 0.1392 / SU Rfree: 0.1272 / Data cutoff high absF: 0 / Data cutoff low absF: 0 / Cross valid method: THROUGHOUT / σ(F): 0 / ESU R: 0.139 / ESU R Free: 0.127 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflectionSelection details
Rfree0.2269 630 4.9 %RANDOM
Rwork0.1964 12320 --
obs0.1979 12950 99.81 %-
Solvent computationIon probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK
Displacement parametersBiso max: 58.16 Å2 / Biso mean: 22.129 Å2 / Biso min: 10.81 Å2
Baniso -1Baniso -2Baniso -3
1-0 Å20 Å2-0.01 Å2
2--0.01 Å20 Å2
3----0.01 Å2
Refinement stepCycle: final / Resolution: 1.8→37.94 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1031 0 26 67 1124
Biso mean--16.76 22.96 -
Num. residues----129
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
X-RAY DIFFRACTIONr_bond_refined_d0.0180.021111
X-RAY DIFFRACTIONr_bond_other_d0.0010.021094
X-RAY DIFFRACTIONr_angle_refined_deg1.82321506
X-RAY DIFFRACTIONr_angle_other_deg0.80632534
X-RAY DIFFRACTIONr_dihedral_angle_1_deg6.4865140
X-RAY DIFFRACTIONr_dihedral_angle_2_deg35.43225.250
X-RAY DIFFRACTIONr_dihedral_angle_3_deg15.24315216
X-RAY DIFFRACTIONr_dihedral_angle_4_deg7.998155
X-RAY DIFFRACTIONr_chiral_restr0.1160.2163
X-RAY DIFFRACTIONr_gen_planes_refined0.0110.021235
X-RAY DIFFRACTIONr_gen_planes_other0.0010.02242
LS refinement shellResolution: 1.8→1.847 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0.295 47 -
Rwork0.26 905 -
all-952 -
obs--99.79 %
Refinement TLS params.

Method: refined / Refine-ID: X-RAY DIFFRACTION

IDL11 (°2)L12 (°2)L13 (°2)L22 (°2)L23 (°2)L33 (°2)S11 (Å °)S12 (Å °)S13 (Å °)S21 (Å °)S22 (Å °)S23 (Å °)S31 (Å °)S32 (Å °)S33 (Å °)T11 (Å2)T12 (Å2)T13 (Å2)T22 (Å2)T23 (Å2)T33 (Å2)Origin x (Å)Origin y (Å)Origin z (Å)
12.235-0.4893-0.14994.48371.14092.059-0.0578-0.0057-0.20530.08060.1311-0.05930.15920.0899-0.07320.07570.022100.07220.01730.050612.99-3.5723.533
22.598-0.9056-0.28634.1014-0.20430.0695-0.0791-0.21060.08180.1590.0991-0.17550.00090.0385-0.020.0590.0162-0.01930.1073-0.02220.045915.1675.5946.355
32.2824-0.2453-0.07972.26430.17061.1218-0.05110.0033-0.03670.01510.06690.12710.0572-0.0053-0.01580.0477-0.0023-0.01620.04240.01660.01768.0934.6163.916
43.1129-0.04641.34753.92441.67625.1879-0.165-0.38130.26180.32620.080.3791-0.248-0.19610.08490.06130.03150.03360.0537-0.0110.0939-1.09915.27911.173
Refinement TLS group
IDRefine-IDRefine TLS-IDAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1A7 - 27
2X-RAY DIFFRACTION2A28 - 53
3X-RAY DIFFRACTION3A54 - 123
4X-RAY DIFFRACTION4A124 - 141

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