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- PDB-6z89: human GTP cyclohydrolase I in complex with allosteric inhibitor -

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Basic information

Entry
Database: PDB / ID: 6z89
Titlehuman GTP cyclohydrolase I in complex with allosteric inhibitor
ComponentsGTP cyclohydrolase 1GTP cyclohydrolase I
KeywordsHYDROLASE / GTP cyclohydrolase I / EC:3.5.4.16 / Tetrahydrobiopterin (BH4) synthesis / Cytosol / Zinc Ion Binding / Hydrolase Activity / Metal Ion Binding / Nucleotide Binding / allosteric inhibitor
Function / homology
Function and homology information


pteridine-containing compound biosynthetic process / regulation of lung blood pressure / GTP cyclohydrolase I / GTP cyclohydrolase I activity / : / neuromuscular process controlling posture / regulation of removal of superoxide radicals / GTP-dependent protein binding / tetrahydrobiopterin biosynthetic process / neuron projection terminus ...pteridine-containing compound biosynthetic process / regulation of lung blood pressure / GTP cyclohydrolase I / GTP cyclohydrolase I activity / : / neuromuscular process controlling posture / regulation of removal of superoxide radicals / GTP-dependent protein binding / tetrahydrobiopterin biosynthetic process / neuron projection terminus / dopamine biosynthetic process / mitogen-activated protein kinase binding / negative regulation of cardiac muscle cell apoptotic process / response to pain / positive regulation of heart rate / response to type II interferon / negative regulation of cellular senescence / response to tumor necrosis factor / Tetrahydrobiopterin (BH4) synthesis, recycling, salvage and regulation / tetrahydrofolate biosynthetic process / positive regulation of telomere maintenance via telomerase / nitric oxide biosynthetic process / negative regulation of blood pressure / positive regulation of nitric-oxide synthase activity / regulation of blood pressure / vasodilation / positive regulation of neuron apoptotic process / cytoplasmic vesicle / protein-containing complex assembly / nuclear membrane / response to lipopolysaccharide / GTPase activity / calcium ion binding / protein-containing complex binding / GTP binding / protein homodimerization activity / protein-containing complex / mitochondrion / zinc ion binding / nucleoplasm / identical protein binding / nucleus / cytosol / cytoplasm
Similarity search - Function
GTP cyclohydrolase I signature 2. / GTP cyclohydrolase I / GTP cyclohydrolase I, conserved site / GTP cyclohydrolase I domain / GTP cyclohydrolase I, N-terminal domain / GTP cyclohydrolase I / GTP cyclohydrolase I signature 1. / GTP cyclohydrolase I, C-terminal/NADPH-dependent 7-cyano-7-deazaguanine reductase
Similarity search - Domain/homology
5-azanyl-[1,3]thiazolo[5,4-d]pyrimidine-2,7-dione / GTP cyclohydrolase 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 2.366 Å
AuthorsEbenhoch, R. / Nar, H.
CitationJournal: Proc Natl Acad Sci U S A / Year: 2020
Title: A hybrid approach reveals the allosteric regulation of GTP cyclohydrolase I.
Authors: Rebecca Ebenhoch / Simone Prinz / Susann Kaltwasser / Deryck J Mills / Robert Meinecke / Martin Rübbelke / Dirk Reinert / Margit Bauer / Lisa Weixler / Markus Zeeb / Janet Vonck / Herbert Nar /
Abstract: Guanosine triphosphate (GTP) cyclohydrolase I (GCH1) catalyzes the conversion of GTP to dihydroneopterin triphosphate (H2NTP), the initiating step in the biosynthesis of tetrahydrobiopterin (BH4). ...Guanosine triphosphate (GTP) cyclohydrolase I (GCH1) catalyzes the conversion of GTP to dihydroneopterin triphosphate (H2NTP), the initiating step in the biosynthesis of tetrahydrobiopterin (BH4). Besides other roles, BH4 functions as cofactor in neurotransmitter biosynthesis. The BH4 biosynthetic pathway and GCH1 have been identified as promising targets to treat pain disorders in patients. The function of mammalian GCH1s is regulated by a metabolic sensing mechanism involving a regulator protein, GCH1 feedback regulatory protein (GFRP). GFRP binds to GCH1 to form inhibited or activated complexes dependent on availability of cofactor ligands, BH4 and phenylalanine, respectively. We determined high-resolution structures of human GCH1-GFRP complexes by cryoelectron microscopy (cryo-EM). Cryo-EM revealed structural flexibility of specific and relevant surface lining loops, which previously was not detected by X-ray crystallography due to crystal packing effects. Further, we studied allosteric regulation of isolated GCH1 by X-ray crystallography. Using the combined structural information, we are able to obtain a comprehensive picture of the mechanism of allosteric regulation. Local rearrangements in the allosteric pocket upon BH4 binding result in drastic changes in the quaternary structure of the enzyme, leading to a more compact, tense form of the inhibited protein, and translocate to the active site, leading to an open, more flexible structure of its surroundings. Inhibition of the enzymatic activity is not a result of hindrance of substrate binding, but rather a consequence of accelerated substrate binding kinetics as shown by saturation transfer difference NMR (STD-NMR) and site-directed mutagenesis. We propose a dissociation rate controlled mechanism of allosteric, noncompetitive inhibition.
History
DepositionJun 2, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0Dec 9, 2020Provider: repository / Type: Initial release
Revision 1.1Dec 23, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.2Jan 24, 2024Group: 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
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: GTP cyclohydrolase 1
B: GTP cyclohydrolase 1
C: GTP cyclohydrolase 1
D: GTP cyclohydrolase 1
E: GTP cyclohydrolase 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)127,31612
Polymers126,6255
Non-polymers6917
Water1,76598
1
A: GTP cyclohydrolase 1
B: GTP cyclohydrolase 1
C: GTP cyclohydrolase 1
D: GTP cyclohydrolase 1
E: GTP cyclohydrolase 1
hetero molecules

A: GTP cyclohydrolase 1
B: GTP cyclohydrolase 1
C: GTP cyclohydrolase 1
D: GTP cyclohydrolase 1
E: GTP cyclohydrolase 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)254,63224
Polymers253,24910
Non-polymers1,38314
Water18010
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
crystal symmetry operation10_555-y,-x,-z+1/61
Buried area33550 Å2
ΔGint-515 kcal/mol
Surface area62270 Å2
MethodPISA
Unit cell
Length a, b, c (Å)121.786, 121.786, 357.15
Angle α, β, γ (deg.)90, 90, 120
Int Tables number179
Space group name H-MP6522

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Components

#1: Protein
GTP cyclohydrolase 1 / GTP cyclohydrolase I / GTP cyclohydrolase I / GTP-CH-I


Mass: 25324.920 Da / Num. of mol.: 5
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GCH1, DYT5, GCH / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P30793, GTP cyclohydrolase I
#2: Chemical ChemComp-QBK / 5-azanyl-[1,3]thiazolo[5,4-d]pyrimidine-2,7-dione


Mass: 182.160 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C5H2N4O2S / Feature type: SUBJECT OF INVESTIGATION
#3: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Zn
#4: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 98 / Source method: isolated from a natural source / Formula: H2O
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: 3.02 Å3/Da / Density % sol: 59.26 %
Crystal growTemperature: 293.15 K / Method: vapor diffusion, sitting drop / pH: 7 / Details: 0.2 M Magnesium chloride, PEG8000; TRIS pH 7.0

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: SLS / Beamline: X06DA / Wavelength: 0.99987 Å
DetectorType: DECTRIS PILATUS 2M-F / Detector: PIXEL / Date: Apr 21, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.99987 Å / Relative weight: 1
ReflectionResolution: 2.366→105.4698 Å / Num. obs: 44678 / % possible obs: 96.1 % / Redundancy: 26.8 % / CC1/2: 1 / Net I/σ(I): 24.8
Reflection shellResolution: 2.366→2.634 Å / Redundancy: 25.3 % / Mean I/σ(I) obs: 1.6 / Num. unique obs: 2235 / CC1/2: 0.686 / % possible all: 78.8

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Processing

Software
NameVersionClassification
BUSTER2.11.7refinement
PDB_EXTRACT3.25data extraction
XDSdata reduction
Aimlessdata scaling
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 1FB1

1fb1


Resolution: 2.366→105.4698 Å / Cor.coef. Fo:Fc: 0.914 / Cor.coef. Fo:Fc free: 0.91 / SU R Cruickshank DPI: 0.376 / Cross valid method: THROUGHOUT / SU R Blow DPI: 0.379 / SU Rfree Blow DPI: 0.243 / SU Rfree Cruickshank DPI: 0.244
RfactorNum. reflection% reflectionSelection details
Rfree0.2378 2220 -RANDOM
Rwork0.2335 ---
obs0.2338 44685 68.3 %-
Displacement parameters
Baniso -1Baniso -2Baniso -3
1-2.9321 Å20 Å20 Å2
2--2.9321 Å20 Å2
3----5.8641 Å2
Refine analyzeLuzzati coordinate error obs: 0.4 Å
Refinement stepCycle: LAST / Resolution: 2.366→105.4698 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms6481 0 29 98 6608
Refine LS restraints
Refine-IDTypeDev idealNumberRestraint functionWeight
X-RAY DIFFRACTIONt_bond_d0.0086623HARMONIC2
X-RAY DIFFRACTIONt_angle_deg1.068943HARMONIC2
X-RAY DIFFRACTIONt_dihedral_angle_d2368SINUSOIDAL2
X-RAY DIFFRACTIONt_gen_planes1104HARMONIC5
X-RAY DIFFRACTIONt_it6623HARMONIC10
X-RAY DIFFRACTIONt_chiral_improper_torsion883SEMIHARMONIC5
X-RAY DIFFRACTIONt_ideal_dist_contact5160SEMIHARMONIC4
X-RAY DIFFRACTIONt_omega_torsion3.24
X-RAY DIFFRACTIONt_other_torsion18.78
LS refinement shellResolution: 2.36→2.54 Å /
RfactorNum. reflection
Rfree0.3385 42
Rwork0.2547 -
Refinement TLS params.

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)
10.4851-0.5122-0.98441.3621.64714.42050.037-0.07940.0356-0.07940.0202-0.07640.0356-0.0764-0.0572-0.0066-0.0286-0.0307-0.0182-0.0116-0.0714-77.554629.726628.548
21.398-0.88152.90831.3635-1.76216.9233-0.0601-0.07750.0022-0.0775-0.0527-0.32360.0022-0.32360.11280.0371-0.0680.0079-0.0490.0604-0.1404-60.5035.22145.2209
36.91013.2026-3.2528.6672-0.6534.32970.2538-0.26750.5447-0.2675-0.2953-0.50020.5447-0.50020.0414-0.07220.10190.0549-0.26150.1012-0.0659-41.83820.613327.2241
42.69553.1232-1.60695.9462-1.29451.6084-0.3471-0.22210.362-0.2221-0.09560.14230.3620.14230.44270.06080.04990.2101-0.14250.011-0.0438-30.049622.12455.8465
51.7937-0.9925-0.00656.3057-0.69750.2035-0.088-0.59520.0083-0.59520.08450.15430.00830.15430.00350.0324-0.0183-0.02120.089-0.0157-0.1816-60.309239.87255.3644
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1{ A|* }A58 - 248
2X-RAY DIFFRACTION2{ B|* }B60 - 249
3X-RAY DIFFRACTION2{ B|* }B251 - 301
4X-RAY DIFFRACTION3{ C|* }C120 - 249
5X-RAY DIFFRACTION3{ C|* }C251
6X-RAY DIFFRACTION4{ D|* }D58 - 249
7X-RAY DIFFRACTION4{ D|* }D251
8X-RAY DIFFRACTION5{ E|* }E58 - 249
9X-RAY DIFFRACTION5{ E|* }E251 - 301

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