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- PDB-7yiv: The Crystal Structure of Human Tissue Nonspecific Alkaline Phosph... -

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Basic information

Entry
Database: PDB / ID: 7yiv
TitleThe Crystal Structure of Human Tissue Nonspecific Alkaline Phosphatase (ALPL) at Basic pH
ComponentsAlkaline phosphatase, tissue-nonspecific isozyme
KeywordsHYDROLASE / alkaline phosphatase / bone mineralization / catalytic network / hypophosphatasia / structural biology
Function / homology
Function and homology information


phosphoamidase / pyridoxal phosphate metabolic process / phosphoamidase activity / phosphoethanolamine phosphatase activity / response to vitamin B6 / futile creatine cycle / pyridoxal phosphatase activity / developmental process involved in reproduction / ADP phosphatase activity / response to macrophage colony-stimulating factor ...phosphoamidase / pyridoxal phosphate metabolic process / phosphoamidase activity / phosphoethanolamine phosphatase activity / response to vitamin B6 / futile creatine cycle / pyridoxal phosphatase activity / developmental process involved in reproduction / ADP phosphatase activity / response to macrophage colony-stimulating factor / inhibition of non-skeletal tissue mineralization / pyrophosphatase activity / Post-translational modification: synthesis of GPI-anchored proteins / cementum mineralization / alkaline phosphatase / alkaline phosphatase activity / response to sodium phosphate / phosphate ion homeostasis / inorganic diphosphate phosphatase activity / cellular homeostasis / endochondral ossification / response to vitamin D / bone mineralization / cellular response to organic cyclic compound / calcium ion homeostasis / side of membrane / dephosphorylation / response to glucocorticoid / extracellular matrix / skeletal system development / mitochondrial membrane / response to insulin / mitochondrial intermembrane space / osteoblast differentiation / positive regulation of cold-induced thermogenesis / response to lipopolysaccharide / response to antibiotic / calcium ion binding / ATP hydrolysis activity / extracellular exosome / extracellular region / membrane / plasma membrane
Similarity search - Function
Alkaline phosphatase, active site / Alkaline phosphatase active site. / Alkaline phosphatase / Alkaline phosphatase / Alkaline phosphatase homologues / Alkaline-phosphatase-like, core domain superfamily
Similarity search - Domain/homology
Alkaline phosphatase, tissue-nonspecific isozyme
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.18 Å
AuthorsCao, Y. / Qin, A. / Yu, Y.T. / Yao, D.Q. / Zhang, Q. / Rao, B. / Xia, Y. / Lu, Y.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Nat Commun / Year: 2023
Title: The structural pathology for hypophosphatasia caused by malfunctional tissue non-specific alkaline phosphatase.
Authors: Yating Yu / Kewei Rong / Deqiang Yao / Qing Zhang / Xiankun Cao / Bing Rao / Ying Xia / Yi Lu / Yafeng Shen / Ying Yao / Hongtao Xu / Peixiang Ma / Yu Cao / An Qin /
Abstract: Hypophosphatasia (HPP) is a metabolic bone disease that manifests as developmental abnormalities in bone and dental tissues. HPP patients exhibit hypo-mineralization and osteopenia due to the ...Hypophosphatasia (HPP) is a metabolic bone disease that manifests as developmental abnormalities in bone and dental tissues. HPP patients exhibit hypo-mineralization and osteopenia due to the deficiency or malfunction of tissue non-specific alkaline phosphatase (TNAP), which catalyzes the hydrolysis of phosphate-containing molecules outside the cells, promoting the deposition of hydroxyapatite in the extracellular matrix. Despite the identification of hundreds of pathogenic TNAP mutations, the detailed molecular pathology of HPP remains unclear. Here, to address this issue, we determine the crystal structures of human TNAP at near-atomic resolution and map the major pathogenic mutations onto the structure. Our study reveals an unexpected octameric architecture for TNAP, which is generated by the tetramerization of dimeric TNAPs, potentially stabilizing the TNAPs in the extracellular environments. Moreover, we use cryo-electron microscopy to demonstrate that the TNAP agonist antibody (JTALP001) forms a stable complex with TNAP by binding to the octameric interface. The administration of JTALP001 enhances osteoblast mineralization and promoted recombinant TNAP-rescued mineralization in TNAP knockout osteoblasts. Our findings elucidate the structural pathology of HPP and highlight the therapeutic potential of the TNAP agonist antibody for osteoblast-associated bone disorders.
History
DepositionJul 18, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jul 19, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 29, 2023Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / pdbx_initial_refinement_model / pdbx_validate_planes
Item: _pdbx_validate_planes.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Alkaline phosphatase, tissue-nonspecific isozyme
D: Alkaline phosphatase, tissue-nonspecific isozyme
C: Alkaline phosphatase, tissue-nonspecific isozyme
B: Alkaline phosphatase, tissue-nonspecific isozyme
G: Alkaline phosphatase, tissue-nonspecific isozyme
H: Alkaline phosphatase, tissue-nonspecific isozyme
F: Alkaline phosphatase, tissue-nonspecific isozyme
E: Alkaline phosphatase, tissue-nonspecific isozyme
hetero molecules


Theoretical massNumber of molelcules
Total (without water)468,05372
Polymers456,1628
Non-polymers11,89164
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area52820 Å2
ΔGint-507 kcal/mol
Surface area130560 Å2
Unit cell
Length a, b, c (Å)158.622, 167.348, 188.783
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121

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Components

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Protein , 1 types, 8 molecules ADCBGHFE

#1: Protein
Alkaline phosphatase, tissue-nonspecific isozyme / / AP-TNAP / TNS-ALP / TNSALP / Alkaline phosphatase liver/bone/kidney isozyme / Phosphoamidase / ...AP-TNAP / TNS-ALP / TNSALP / Alkaline phosphatase liver/bone/kidney isozyme / Phosphoamidase / Phosphocreatine phosphatase


Mass: 57020.188 Da / Num. of mol.: 8
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ALPL / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P05186, alkaline phosphatase, phosphoamidase

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Sugars , 2 types, 32 molecules

#2: Polysaccharide
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 16
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#3: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 3 types, 32 molecules

#4: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#5: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: Zn / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 8 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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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: 2.75 Å3/Da / Density % sol: 55.21 %
Crystal growTemperature: 293 K / Method: vapor diffusion, sitting drop / pH: 8
Details: 17% (w/v) PEG4000, 30 mM Tris, pH 8.0, 100 mM MgCl2

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Data collection

DiffractionMean temperature: 77 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SSRF / Beamline: BL18U1 / Wavelength: 1 Å
DetectorType: DECTRIS PILATUS3 6M / Detector: PIXEL / Date: Jan 31, 2021
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 3.17→43.49 Å / Num. obs: 83898 / % possible obs: 98.62 % / Redundancy: 6.7 % / Biso Wilson estimate: 70.29 Å2 / CC1/2: 0.98 / Net I/σ(I): 2.28
Reflection shellResolution: 3.2→3.26 Å / Redundancy: 6.5 % / Num. unique obs: 11103 / CC1/2: 0.663 / % possible all: 98.5

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Processing

Software
NameVersionClassification
PHENIX1.19.2_4158refinement
xia2data scaling
PDB_EXTRACT3.27data extraction
HKL-2000data reduction
PHENIXphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 1EW2

1ew2


Resolution: 3.18→25.05 Å / SU ML: 0.4 / Cross valid method: THROUGHOUT / σ(F): 1.34 / Phase error: 24.36 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.2339 1993 2.38 %
Rwork0.1723 81905 -
obs0.1737 83898 98.62 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Displacement parametersBiso max: 137.86 Å2 / Biso mean: 68.6681 Å2 / Biso min: 28.54 Å2
Refinement stepCycle: final / Resolution: 3.18→25.05 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms29969 0 704 0 30673
Biso mean--103.51 --
Num. residues----3869
LS refinement shell

Refine-ID: X-RAY DIFFRACTION / Rfactor Rfree error: 0 / Total num. of bins used: 14

Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkNum. reflection all% reflection obs (%)
3.18-3.260.3021320.24625187531989
3.26-3.340.31841310.24025794592599
3.34-3.440.31641470.24665800594799
3.44-3.550.31651340.22865853598799
3.55-3.680.2891490.20945823597299
3.68-3.830.24591480.19615841598999
3.83-40.28361400.20145859599999
4-4.210.25641360.161858856021100
4.21-4.470.18291440.141958826026100
4.47-4.810.18351450.128559296074100
4.81-5.290.19971400.140659246064100
5.29-6.050.2131470.155459556102100
6.05-7.590.21391440.164260156159100
7.59-25.050.19821560.14996158631499

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