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Yorodumi- PDB-6hyc: The structure of full-length human phenylalanine hydroxylase in c... -
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-Basic information
Entry | Database: PDB / ID: 6hyc | |||||||||
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Title | The structure of full-length human phenylalanine hydroxylase in complex with the cofactor and negative regulator tetrahydrobiopterin | |||||||||
Components | Phenylalanine-4-hydroxylase | |||||||||
Keywords | OXIDOREDUCTASE / Tetrahydrobiopterin / phenylalanine hydroxylase / phenylketonuria / allostery | |||||||||
Function / homology | Function and homology information Phenylketonuria / Phenylalanine metabolism / phenylalanine 4-monooxygenase / phenylalanine 4-monooxygenase activity / tyrosine biosynthetic process / catecholamine biosynthetic process / L-phenylalanine catabolic process / amino acid biosynthetic process / iron ion binding / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | X-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.18 Å | |||||||||
Authors | Alcorlo Pages, M. / Flydal, I.M. | |||||||||
Funding support | Spain, Norway, 2items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2019 Title: Structure of full-length human phenylalanine hydroxylase in complex with tetrahydrobiopterin. Authors: Marte Innselset Flydal / Martín Alcorlo-Pagés / Fredrik Gullaksen Johannessen / Siseth Martínez-Caballero / Lars Skjærven / Rafael Fernandez-Leiro / Aurora Martinez / Juan A Hermoso / Abstract: Phenylalanine hydroxylase (PAH) is a key enzyme in the catabolism of phenylalanine, and mutations in this enzyme cause phenylketonuria (PKU), a genetic disorder that leads to brain damage and mental ...Phenylalanine hydroxylase (PAH) is a key enzyme in the catabolism of phenylalanine, and mutations in this enzyme cause phenylketonuria (PKU), a genetic disorder that leads to brain damage and mental retardation if untreated. Some patients benefit from supplementation with a synthetic formulation of the cofactor tetrahydrobiopterin (BH) that partly acts as a pharmacological chaperone. Here we present structures of full-length human PAH (hPAH) both unbound and complexed with BH in the precatalytic state. Crystal structures, solved at 3.18-Å resolution, show the interactions between the cofactor and PAH, explaining the negative regulation exerted by BH BH forms several H-bonds with the N-terminal autoregulatory tail but is far from the catalytic Fe Upon BH binding a polar and salt-bridge interaction network links the three PAH domains, explaining the stability conferred by BH Importantly, BH binding modulates the interaction between subunits, providing information about PAH allostery. Moreover, we also show that the cryo-EM structure of hPAH in absence of BH reveals a highly dynamic conformation for the tetramers. Structural analyses of the hPAH:BH subunits revealed that the substrate-induced movement of Tyr138 into the active site could be coupled to the displacement of BH from the precatalytic toward the active conformation, a molecular mechanism that was supported by site-directed mutagenesis and targeted molecular dynamics simulations. Finally, comparison of the rat and human PAH structures show that hPAH is more dynamic, which is related to amino acid substitutions that enhance the flexibility of hPAH and may increase the susceptibility to PKU-associated mutations. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 6hyc.cif.gz | 348 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6hyc.ent.gz | 284.3 KB | Display | PDB format |
PDBx/mmJSON format | 6hyc.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/hy/6hyc ftp://data.pdbj.org/pub/pdb/validation_reports/hy/6hyc | HTTPS FTP |
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-Related structure data
Related structure data | 4605C 6hpoC 5denS S: Starting model for refinement C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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2 |
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Unit cell |
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-Components
#1: Protein | Mass: 51928.906 Da / Num. of mol.: 4 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: PAH / Production host: Escherichia coli #1/H766 (bacteria) / References: UniProt: P00439, phenylalanine 4-monooxygenase #2: Chemical | #3: Water | ChemComp-HOH / | |
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-Experimental details
-Experiment
Experiment | Method: X-RAY DIFFRACTION / Number of used crystals: 1 |
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-Sample preparation
Crystal | Density Matthews: 2.48 Å3/Da / Density % sol: 50.35 % |
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Crystal grow | Temperature: 291 K / Method: vapor diffusion, sitting drop / pH: 7 Details: 1.5 M DL Malic Acid pH=7.0, 100 mM Bis-Tris Propane pH=6.2, 0.9 mM Thesit, 0.98 mM BH4, 25 mM DTT and 1 mM reduced glutathione |
-Data collection
Diffraction | Mean temperature: 100 K / Serial crystal experiment: N |
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Diffraction source | Source: SYNCHROTRON / Site: ALBA / Beamline: XALOC / Wavelength: 0.97926 Å |
Detector | Type: DECTRIS PILATUS3 S 6M / Detector: PIXEL / Date: Oct 19, 2016 |
Radiation | Protocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray |
Radiation wavelength | Wavelength: 0.97926 Å / Relative weight: 1 |
Reflection | Resolution: 3.18→33.92 Å / Num. obs: 34280 / % possible obs: 94.3 % / Redundancy: 3 % / Rmerge(I) obs: 0.181 / Rpim(I) all: 0.1314 / Net I/σ(I): 2.4 |
Reflection shell | Resolution: 3.18→3.34 Å / Num. unique obs: 3926 |
-Processing
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Refinement | Method to determine structure: MOLECULAR REPLACEMENT Starting model: 5den Resolution: 3.18→33.92 Å / SU ML: 0.43 / Cross valid method: FREE R-VALUE / σ(F): 0 / Phase error: 31.58
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Solvent computation | Shrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement step | Cycle: LAST / Resolution: 3.18→33.92 Å
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Refine LS restraints |
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LS refinement shell |
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