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- PDB-2aut: Crystal structure of Lys154Asn mutant of mature AphA of S. typhimurium -

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Basic information

Entry
Database: PDB / ID: 2aut
TitleCrystal structure of Lys154Asn mutant of mature AphA of S. typhimurium
ComponentsAphA
KeywordsHYDROLASE / Class-B bacterial non-specific acid phosphatase / Lys154Asn mutant of mature AphA / metalloenzyme
Function / homology
Function and homology information


acid phosphatase / acid phosphatase activity / outer membrane-bounded periplasmic space / metal ion binding
Similarity search - Function
HAD-superfamily phosphatase, subfamily IIIB, AphA / Acid phosphatase, class B-like / HAD superfamily, subfamily IIIB (Acid phosphatase) / HAD superfamily/HAD-like / HAD superfamily / HAD-like superfamily / Rossmann fold / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
PHOSPHATE ION / Class B acid phosphatase / Class B acid phosphatase
Similarity search - Component
Biological speciesSalmonella typhimurium (bacteria)
MethodX-RAY DIFFRACTION / FOURIER SYNTHESIS / Resolution: 2.25 Å
AuthorsMakde, R.D. / Gupta, G.D. / Kumar, V.
Citation
Journal: Arch.Biochem.Biophys. / Year: 2007
Title: Structural and mutational analyses reveal the functional role of active-site Lys-154 and Asp-173 of Salmonella typhimurium AphA protein.
Authors: Makde, R.D. / Gupta, G.D. / Mahajan, S.K. / Kumar, V.
#1: Journal: Acta Crystallogr.,Sect.D / Year: 2003
Title: Expression, purification, crystallization and preliminary X-ray diffraction studies of recombinant class B non-specific acid phosphatase of Salmonella typhimurium
Authors: Makde, R.D. / Kumar, V. / Gupta, G.D. / Jasti, J. / Singh, T.P. / Mahajan, S.K.
History
DepositionAug 29, 2005Deposition site: RCSB / Processing site: RCSB
Revision 1.0Sep 5, 2006Provider: repository / Type: Initial release
Revision 1.1May 1, 2008Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Version format compliance
Revision 1.3Oct 11, 2017Group: Refinement description / Category: software / Item: _software.name
Revision 1.4Oct 20, 2021Group: Database references / Derived calculations
Category: database_2 / pdbx_struct_conn_angle ...database_2 / 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_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.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_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_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
Revision 1.5Aug 23, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: AphA
B: AphA
C: AphA
D: AphA
hetero molecules


Theoretical massNumber of molelcules
Total (without water)95,63810
Polymers95,4234
Non-polymers2156
Water9,620534
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area12330 Å2
ΔGint-128 kcal/mol
Surface area34450 Å2
MethodPISA
Unit cell
Length a, b, c (Å)70.737, 84.477, 149.268
Angle α, β, γ (deg.)90.00, 90.00, 90.00
Int Tables number19
Space group name H-MP212121
DetailsThe entry contains the crystallographic asymmetric unit consisting of four chains (homotetramer), which is the known biologically active state of the APHA protein.

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Components

#1: Protein
AphA /


Mass: 23855.625 Da / Num. of mol.: 4 / Mutation: K154N
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Salmonella typhimurium (bacteria) / Gene: aphA / Plasmid: pET21(A) / Species (production host): Escherichia coli / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3)
References: UniProt: P58683, UniProt: Q540U1*PLUS, acid phosphatase
#2: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Mg
#3: Chemical ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Na
#4: Chemical ChemComp-PO4 / PHOSPHATE ION / Phosphate


Mass: 94.971 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: PO4
#5: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 534 / 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: 2.34 Å3/Da / Density % sol: 47.4 %
Crystal growTemperature: 293 K / Method: vapor diffusion sitting-drop, micro-seeding / pH: 4.7
Details: PEG 6000, magnesium chloride, sodium acetate, dihydrogen phosphate , pH 4.7, vapour diffusion sitting-drop, micro-seeding, temperature 293K

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Data collection

DiffractionMean temperature: 293 K
Diffraction sourceSource: ROTATING ANODE / Type: RIGAKU RU200 / Wavelength: 1.5418 Å
DetectorType: MARRESEARCH / Detector: IMAGE PLATE / Date: Jan 31, 2004 / Details: OSMIC mirror
RadiationMonochromator: GRAPHITE / Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.5418 Å / Relative weight: 1
ReflectionResolution: 2.24→30 Å / Num. all: 40809 / Num. obs: 40671 / % possible obs: 94.5 % / Observed criterion σ(I): 0 / Redundancy: 3.4 % / Biso Wilson estimate: 24.77 Å2 / Rmerge(I) obs: 0.055 / Net I/σ(I): 13.5
Reflection shellResolution: 2.24→2.36 Å / Redundancy: 3.2 % / Rmerge(I) obs: 0.162 / Mean I/σ(I) obs: 7.1 / Num. unique all: 5241 / % possible all: 91.4

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Processing

Software
NameVersionClassification
MOSFLMdata reduction
SCALA/ TRUNCATEdata scaling
CNSrefinement
CCP4(SCALAdata scaling
TRUNCATEdata scaling
CNSphasing
RefinementMethod to determine structure: FOURIER SYNTHESIS
Starting model: Partially refined structure of wild-type APHA (PDB code 1Z5G) with all HET atoms (HOH, MG etc.) removed and side chain of Lys154 replaced with ALA was used as the starting model. A ...Starting model: Partially refined structure of wild-type APHA (PDB code 1Z5G) with all HET atoms (HOH, MG etc.) removed and side chain of Lys154 replaced with ALA was used as the starting model. A random error of 0.2A was added to the atomic coordinates of the starting model.

1z5g


Resolution: 2.25→20 Å / Isotropic thermal model: ANISOTROPIC / Cross valid method: THROUGHOUT / σ(F): 0 / Stereochemistry target values: Engh & Huber
RfactorNum. reflection% reflectionSelection details
Rfree0.197 2004 5 %random
Rwork0.151 ---
all-40554 --
obs-40554 94 %-
Solvent computationSolvent model: Flat Model / Bsol: 31.75 Å2 / ksol: 0.327 e/Å3
Displacement parametersBiso mean: 24.29 Å2
Baniso -1Baniso -2Baniso -3
1-0.142 Å20 Å20 Å2
2---4.802 Å20 Å2
3---4.659 Å2
Refine analyze
FreeObs
Luzzati coordinate error0.25 Å0.19 Å
Luzzati d res low-5 Å
Luzzati sigma a0.21 Å0.16 Å
Refinement stepCycle: LAST / Resolution: 2.25→20 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6589 0 10 534 7133
Refine LS restraints
Refine-IDTypeDev idealDev ideal target
X-RAY DIFFRACTIONc_bond_d0.006
X-RAY DIFFRACTIONc_angle_deg1
X-RAY DIFFRACTIONc_dihedral_angle_d23.09
X-RAY DIFFRACTIONc_improper_angle_d0.833
X-RAY DIFFRACTIONc_mcbond_it1.41.5
X-RAY DIFFRACTIONc_mcangle_it2.222
X-RAY DIFFRACTIONc_scbond_it2.172
X-RAY DIFFRACTIONc_scangle_it3.22.5
LS refinement shellResolution: 2.25→2.33 Å / Total num. of bins used: 10
RfactorNum. reflection% reflection
Rfree0.228 192 5 %
Rwork0.174 3672 -
obs-3864 91 %
Xplor file
Refine-IDSerial noParam fileTopol file
X-RAY DIFFRACTION1PROTEIN_REP.PARAMPROTEIN.TOP
X-RAY DIFFRACTION2WATER_REP.PARAMWATER.TOP
X-RAY DIFFRACTION3ION.PARAMION.TOP

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