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- PDB-1ew8: ALKALINE PHOSPHATASE (E.C. 3.1.3.1) COMPLEX WITH PHOSPHONOACETIC ACID -

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Basic information

Entry
Database: PDB / ID: 1ew8
TitleALKALINE PHOSPHATASE (E.C. 3.1.3.1) COMPLEX WITH PHOSPHONOACETIC ACID
ComponentsALKALINE PHOSPHATASE
KeywordsHYDROLASE / enzyme-inhibitor complex
Function / homology
Function and homology information


oxidoreductase activity, acting on phosphorus or arsenic in donors / alkaline phosphatase / alkaline phosphatase activity / hydrogenase (acceptor) activity / phosphoprotein phosphatase activity / dephosphorylation / protein dephosphorylation / outer membrane-bounded periplasmic space / periplasmic space / magnesium ion binding / zinc ion binding
Similarity search - Function
Alkaline phosphatase, active site / Alkaline phosphatase active site. / Alkaline phosphatase / Alkaline phosphatase / Alkaline phosphatase homologues / Alkaline Phosphatase, subunit A / Alkaline Phosphatase, subunit A / Alkaline-phosphatase-like, core domain superfamily / 3-Layer(aba) Sandwich / Alpha Beta
Similarity search - Domain/homology
PHOSPHONOACETIC ACID / PHOSPHATE ION / Alkaline phosphatase
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
MethodX-RAY DIFFRACTION / Resolution: 2.2 Å
AuthorsHoltz, K.M. / Stec, B. / Myers, J.K. / Antonelli, S.M. / Widlanski, T.S. / Kantrowitz, E.R.
Citation
Journal: Protein Sci. / Year: 2000
Title: Alternate modes of binding in two crystal structures of alkaline phosphatase-inhibitor complexes.
Authors: Holtz, K.M. / Stec, B. / Myers, J.K. / Antonelli, S.M. / Widlanski, T.S. / Kantrowitz, E.R.
#1: Journal: J.Biol.Chem. / Year: 1999
Title: A Model of the Transition State in the Alkaline Phosphatase Reaction
Authors: Holtz, K.M. / Stec, B. / Kantrowitz, E.R.
#2: Journal: J.Mol.Biol. / Year: 1991
Title: Reaction Mechanism of Alkaline Phosphatase Based on Crystal Structures: Two-metal Ion Catalysis
Authors: Kim, E.E. / Wyckoff, H.W.
History
DepositionApr 24, 2000Deposition site: RCSB / Processing site: RCSB
Revision 1.0May 1, 2002Provider: repository / Type: Initial release
Revision 1.1Oct 16, 2007Group: Version format compliance
Revision 1.2Jul 13, 2011Group: Derived calculations / Version format compliance
Revision 1.3Mar 13, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Source and taxonomy / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / entity / pdbx_entity_src_syn / pdbx_struct_conn_angle / struct_conn / struct_site
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _entity.details / _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_ptnr1_label_alt_id / _struct_conn.pdbx_ptnr2_label_alt_id / _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_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

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Assembly

Deposited unit
A: ALKALINE PHOSPHATASE
B: ALKALINE PHOSPHATASE
hetero molecules


Theoretical massNumber of molelcules
Total (without water)95,29216
Polymers94,1892
Non-polymers1,10314
Water9,314517
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area9950 Å2
ΔGint-343 kcal/mol
Surface area27500 Å2
MethodPISA, PQS
Unit cell
Length a, b, c (Å)194.77, 167.19, 76.59
Angle α, β, γ (deg.)90, 90, 90
Int Tables number23
Space group name H-MI222
Detailshomodimeric metalloenzyme with a non-crystallographic 2-fold symmetry axis

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Components

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Protein , 1 types, 2 molecules AB

#1: Protein ALKALINE PHOSPHATASE /


Mass: 47094.398 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli) / Strain: SM547 / Cellular location: PERIPLASM / Gene: PHOA / Plasmid: PEK154 / Production host: Escherichia coli (E. coli) / References: UniProt: P00634, alkaline phosphatase

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Non-polymers , 6 types, 531 molecules

#2: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Zn / Details: Obtained from Sigma Chemical Co.
#3: Chemical ChemComp-PO4 / PHOSPHATE ION / Phosphate


Mass: 94.971 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: PO4
#4: Chemical ChemComp-SO4 / SULFATE ION / Sulfate


Mass: 96.063 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: SO4
#5: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#6: Chemical ChemComp-PAE / PHOSPHONOACETIC ACID


Mass: 140.032 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C2H5O5P
#7: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 517 / Source method: isolated from a natural source / Formula: H2O

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Details

Nonpolymer detailsThe active site has mixed occupancy with some phosphonoacetic acid (PAE) and some phosphate (PO4). ...The active site has mixed occupancy with some phosphonoacetic acid (PAE) and some phosphate (PO4). The PAE and PO4 are assigned different residue numbers, 556 and 557, and alternate conformation indicators A and B, respectively. The alternate position indicator signifies that these residues occupy the same area in space. The magnesium 452 has three waters associated with it and it has been labelled as ligand MO3 for each chain. The zinc ion 452 has the same three waters associated with it and is labelled ZO3. The ZO3 and MO3 occupy the same space and have the same coordinates, but their occupancies are different. The MO3 is labelled as residue 452, conformation A. The ZO3 is labelled as residue 453, conformation B. SO4 558 is associated with chain A, and SO4 568 is associated with chain B.

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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: 3.31 Å3/Da / Density % sol: 62.82 %
Crystal growTemperature: 298 K / Method: vapor diffusion, hanging drop / pH: 9.5
Details: enzyme: 30 mg/mL; buffer: 40% saturating ammonium sulfate/100 mM Tris/10 mM MgSO4, pH 9.5, VAPOR DIFFUSION, HANGING DROP, temperature 298K
Crystal grow
*PLUS
Components of the solutions
*PLUS
IDConc.Common nameCrystal-IDSol-IDChemical formulaDetails
130 mg/mlenzyme1drop
220 %satammonium sulfate1reservoir
3100 mMTris1reservoir
410 mM1reservoirMgCl2
50.01 mM1reservoirZnCl2pH9.5

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Data collection

DiffractionMean temperature: 295 K
Diffraction sourceSource: ROTATING ANODE / Type: RIGAKU RU200 / Wavelength: 1.5418
DetectorType: UCSD MARK III / Detector: AREA DETECTOR / Date: Jun 5, 1996
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.5418 Å / Relative weight: 1
ReflectionResolution: 2.2→30 Å / Num. all: 63777 / Num. obs: 59057 / % possible obs: 92.6 % / Observed criterion σ(F): 0 / Observed criterion σ(I): 0 / Redundancy: 3.1 % / Biso Wilson estimate: 31.9 Å2 / Rmerge(I) obs: 0.078 / Net I/σ(I): 6.6
Reflection shellResolution: 2.2→2.37 Å / Redundancy: 1.92 % / Rmerge(I) obs: 0.28 / Num. unique all: 11823 / % possible all: 94
Reflection
*PLUS
Num. measured all: 180502
Reflection shell
*PLUS
Lowest resolution: 2.4 Å / % possible obs: 79.6 % / Redundancy: 2 % / Mean I/σ(I) obs: 1.2

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Processing

Software
NameClassification
SDMSdata collection
SDMSdata reduction
X-PLORmodel building
SHELXL-97refinement
SDMSdata scaling
X-PLORphasing
RefinementResolution: 2.2→12 Å / σ(F): 0 / σ(I): 0 / Stereochemistry target values: Engh & Huber
Details: Used conjugated gradient least squares for the refinement and weighted full matrix least squares procedure for estimating ESDs.
RfactorNum. reflection% reflectionSelection details
Rfree0.257 2311 -every 25
Rwork0.189 ---
all0.196 58257 --
obs0.196 58051 99.6 %-
Refinement stepCycle: LAST / Resolution: 2.2→12 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6608 0 44 517 7169
Refine LS restraints
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONs_bond_d0.015
X-RAY DIFFRACTIONs_angle_d0.03
Software
*PLUS
Name: SHELXL / Version: 97 / Classification: refinement
Refinement
*PLUS
% reflection Rfree: 10 % / Rfactor all: 0.19 / Rfactor Rwork: 0.19
Solvent computation
*PLUS
Displacement parameters
*PLUS
Refine LS restraints
*PLUS
Refine-IDTypeDev ideal
X-RAY DIFFRACTIONs_bond_d0.02
X-RAY DIFFRACTIONs_angle_d
X-RAY DIFFRACTIONs_angle_deg1.92
X-RAY DIFFRACTIONs_dihedral_angle_d
X-RAY DIFFRACTIONs_dihedral_angle_deg24.7
X-RAY DIFFRACTIONs_improper_angle_d
X-RAY DIFFRACTIONs_improper_angle_deg1.82

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