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- EMDB-11114: inhibitory human GTP cyclohydrolase I - GFRP complex -

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Basic information

Entry
Database: EMDB / ID: EMD-11114
Titleinhibitory human GTP cyclohydrolase I - GFRP complex
Map data
Sample
  • Complex: inhibitory GCH1-GFRP complex (BH4 bound)
    • Protein or peptide: GTP cyclohydrolase 1
    • Protein or peptide: GTP cyclohydrolase 1 feedback regulatory protein
  • Ligand: ZINC ION
  • Ligand: 7,8-DIHYDROBIOPTERIN
KeywordsGTP cyclohydrolase GFTP / I / EC:3.5.4.16 / Tetrahydrobiopterin (BH4) synthesis / Cytosol / Zinc Ion Binding / Hydrolase Activity / Metal Ion Binding / Nucleotide Binding / allosteric inhibited / HYDROLASE
Function / homology
Function and homology information


GTP cyclohydrolase binding / pteridine-containing compound biosynthetic process / dihydrobiopterin metabolic process / regulation of lung blood pressure / GTP cyclohydrolase I / GTP cyclohydrolase I activity / negative regulation of biosynthetic process / neuromuscular process controlling posture / GTP-dependent protein binding / regulation of removal of superoxide radicals ...GTP cyclohydrolase binding / pteridine-containing compound biosynthetic process / dihydrobiopterin metabolic process / regulation of lung blood pressure / GTP cyclohydrolase I / GTP cyclohydrolase I activity / negative regulation of biosynthetic process / neuromuscular process controlling posture / GTP-dependent protein binding / regulation of removal of superoxide radicals / tetrahydrobiopterin biosynthetic process / neuron projection terminus / regulation of nitric oxide biosynthetic process / mitogen-activated protein kinase binding / dopamine biosynthetic process / negative regulation of cardiac muscle cell apoptotic process / positive regulation of heart rate / response to pain / 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 / melanosome / cytoplasmic vesicle / protein-containing complex assembly / nuclear membrane / response to lipopolysaccharide / GTPase activity / dendrite / 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, feedback regulatory protein / GFRP superfamily / GTP cyclohydrolase I feedback regulatory protein (GFRP) / 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
GTP cyclohydrolase 1 feedback regulatory protein / GTP cyclohydrolase 1
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.9 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, 2020-
Header (metadata) releaseDec 9, 2020-
Map releaseDec 9, 2020-
UpdateMay 22, 2024-
Current statusMay 22, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.05
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  • Surface view with fitted model
  • Atomic models: PDB-6z85
  • Surface level: 0.05
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_11114.png

Downloads & links

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Map

FileDownload / File: emd_11114.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)X (Row.)Y (Col.)
1.08 Å/pix.
x 240 pix.
= 258.48 Å		1.08 Å/pix.
x 240 pix.
= 258.48 Å		1.08 Å/pix.
x 240 pix.
= 258.48 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.077 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.05 / Movie #1: 0.05
Minimum - Maximum-0.3683759 - 0.59016687
Average (Standard dev.)0.00018904875 (±0.012547049)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderYXZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 258.48 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0771.0771.077
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z258.480258.480258.480
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ240240240
MAP C/R/S213
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.3680.5900.000

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Supplemental data

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Sample components

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Entire : inhibitory GCH1-GFRP complex (BH4 bound)

EntireName: inhibitory GCH1-GFRP complex (BH4 bound)
Components
  • Complex: inhibitory GCH1-GFRP complex (BH4 bound)
    • Protein or peptide: GTP cyclohydrolase 1
    • Protein or peptide: GTP cyclohydrolase 1 feedback regulatory protein
  • Ligand: ZINC ION
  • Ligand: 7,8-DIHYDROBIOPTERIN

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Supramolecule #1: inhibitory GCH1-GFRP complex (BH4 bound)

SupramoleculeName: inhibitory GCH1-GFRP complex (BH4 bound) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 353 KDa

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Macromolecule #1: GTP cyclohydrolase 1

MacromoleculeName: GTP cyclohydrolase 1 / type: protein_or_peptide / ID: 1 / Number of copies: 10 / Enantiomer: LEVO / EC number: GTP cyclohydrolase I
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 25.32492 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString: MHHHHHHGSD DDDKRPEAKS AQPADGWKGE RPRSEEDNEL NLPNLAAAYS SILSSLGENP QRQGLLKTPW RAASAMQFFT KGYQETISD VLNDAIFDED HDEMVIVKDI DMFSMCEHHL VPFVGKVHIG YLPNKQVLGL SKLARIVEIY SRRLQVQERL T KQIAVAIT ...String:
MHHHHHHGSD DDDKRPEAKS AQPADGWKGE RPRSEEDNEL NLPNLAAAYS SILSSLGENP QRQGLLKTPW RAASAMQFFT KGYQETISD VLNDAIFDED HDEMVIVKDI DMFSMCEHHL VPFVGKVHIG YLPNKQVLGL SKLARIVEIY SRRLQVQERL T KQIAVAIT EALRPAGVGV VVEATHMCMV MRGVQKMNSK TVTSTMLGVF REDPKTREEF LTLIRS

UniProtKB: GTP cyclohydrolase 1

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Macromolecule #2: GTP cyclohydrolase 1 feedback regulatory protein

MacromoleculeName: GTP cyclohydrolase 1 feedback regulatory protein / type: protein_or_peptide / ID: 2 / Number of copies: 10 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 9.992483 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
GSHMPYLLIS TQIRMEVGPT MVGDEQSDPE LMQHLGASKR RALGNNFYEY YVDDPPRIVL DKLERRGFRV LSMTGVGQTL VWCLHKE

UniProtKB: GTP cyclohydrolase 1 feedback regulatory protein

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Macromolecule #3: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 3 / Number of copies: 10 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #4: 7,8-DIHYDROBIOPTERIN

MacromoleculeName: 7,8-DIHYDROBIOPTERIN / type: ligand / ID: 4 / Number of copies: 10 / Formula: HBI
Molecular weightTheoretical: 239.231 Da
Chemical component information

ChemComp-HBI:
7,8-DIHYDROBIOPTERIN

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.25 mg/mL
BufferpH: 5.75
Component:
ConcentrationFormulaName
40.0 mMNa3C6H5O7sodium citrate
0.1 mMC9H15N5O3Tetrahydrobiopterin
GridModel: C-flat-1.2/1.3 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 2688 / Average exposure time: 8.0 sec. / Average electron dose: 55.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1871460
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

1wpl

Final reconstructionApplied symmetry - Point group: D5 (2x5 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 560802
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.0)
Final 3D classificationSoftware - Name: RELION (ver. 3.0)

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Atomic model buiding 1

Initial model
PDB IDChain

1fb1

source_name: PDB, initial_model_type: experimental model

1wpl

source_name: PDB, initial_model_type: experimental model
RefinementProtocol: RIGID BODY FIT
Output model

PDB-6z85:
inhibitory human GTP cyclohydrolase I - GFRP complex

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