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- PDB-8t1w: Crystal structure of orphan G protein-coupled receptor 6 with bou... -

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Basic information

Entry
Database: PDB / ID: 8t1w
TitleCrystal structure of orphan G protein-coupled receptor 6 with bound CVN424
ComponentsG-protein coupled receptor 6, Soluble cytochrome b562 chimera
KeywordsMEMBRANE PROTEIN / Orphan GPCR / GPR6 / BRIL / CVN424 / inverse agonist / LCP / synchrotron / APS
Function / homology
Function and homology information


sphingosine-1-phosphate receptor activity / regulation of metabolic process / G protein-coupled receptor activity / electron transport chain / adenylate cyclase-activating G protein-coupled receptor signaling pathway / positive regulation of cytosolic calcium ion concentration / periplasmic space / electron transfer activity / G protein-coupled receptor signaling pathway / iron ion binding ...sphingosine-1-phosphate receptor activity / regulation of metabolic process / G protein-coupled receptor activity / electron transport chain / adenylate cyclase-activating G protein-coupled receptor signaling pathway / positive regulation of cytosolic calcium ion concentration / periplasmic space / electron transfer activity / G protein-coupled receptor signaling pathway / iron ion binding / heme binding / plasma membrane / cytoplasm
Similarity search - Function
G protein-coupled receptor 6 / G protein-coupled receptor 3/6/12 orphan / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family)
Similarity search - Domain/homology
: / Soluble cytochrome b562 / G-protein coupled receptor 6
Similarity search - Component
Biological speciesHomo sapiens (human)
Escherichia coli (E. coli)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.49 Å
AuthorsBarekatain, M. / Johansson, L. / Lam, J.H. / Sadybekov, A.V. / Han, G.W. / Popov, P. / Russo, J. / Bliesath, J. / Brice, N. / Beresford, M. ...Barekatain, M. / Johansson, L. / Lam, J.H. / Sadybekov, A.V. / Han, G.W. / Popov, P. / Russo, J. / Bliesath, J. / Brice, N. / Beresford, M. / Carlson, L. / Saikatendu, K.S. / Sun, H. / Murphy, S. / Monenschein, H. / Schiffer, H.H. / Lutomski, C. / Robinson, C.V. / Liu, Z. / Hua, T. / Katritch, V. / Cherezov, V.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM127086 United States
CitationJournal: Sci Signal / Year: 2024
Title: Structural insights into the high basal activity and inverse agonism of the orphan receptor GPR6 implicated in Parkinson's disease.
Authors: Mahta Barekatain / Linda C Johansson / Jordy H Lam / Hao Chang / Anastasiia V Sadybekov / Gye Won Han / Joseph Russo / Joshua Bliesath / Nicola L Brice / Mark B L Carlton / Kumar S ...Authors: Mahta Barekatain / Linda C Johansson / Jordy H Lam / Hao Chang / Anastasiia V Sadybekov / Gye Won Han / Joseph Russo / Joshua Bliesath / Nicola L Brice / Mark B L Carlton / Kumar S Saikatendu / Hukai Sun / Sean T Murphy / Holger Monenschein / Hans H Schiffer / Petr Popov / Corinne A Lutomski / Carol V Robinson / Zhi-Jie Liu / Tian Hua / Vsevolod Katritch / Vadim Cherezov /
Abstract: GPR6 is an orphan G protein-coupled receptor with high constitutive activity found in D2-type dopamine receptor-expressing medium spiny neurons of the striatopallidal pathway, which is aberrantly ...GPR6 is an orphan G protein-coupled receptor with high constitutive activity found in D2-type dopamine receptor-expressing medium spiny neurons of the striatopallidal pathway, which is aberrantly hyperactivated in Parkinson's disease. Here, we solved crystal structures of GPR6 without the addition of a ligand (a pseudo-apo state) and in complex with two inverse agonists, including CVN424, which improved motor symptoms in patients with Parkinson's disease in clinical trials. In addition, we obtained a cryo-electron microscopy structure of the signaling complex between GPR6 and its cognate G heterotrimer. The pseudo-apo structure revealed a strong density in the orthosteric pocket of GPR6 corresponding to a lipid-like endogenous ligand. A combination of site-directed mutagenesis, native mass spectrometry, and computer modeling suggested potential mechanisms for high constitutive activity and inverse agonism in GPR6 and identified a series of lipids and ions bound to the receptor. The structures and results obtained in this study could guide the rational design of drugs that modulate GPR6 signaling.
History
DepositionJun 4, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Dec 4, 2024Provider: repository / Type: Initial release
Revision 1.1Dec 18, 2024Group: Data collection / Database references / Derived calculations
Category: citation / citation_author ...citation / citation_author / pdbx_struct_assembly / pdbx_struct_assembly_gen / pdbx_struct_assembly_prop / pdbx_struct_oper_list / struct_biol
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID / _citation_author.name / _pdbx_struct_assembly.details / _pdbx_struct_assembly.method_details / _pdbx_struct_assembly.oligomeric_count / _pdbx_struct_assembly.oligomeric_details / _pdbx_struct_assembly_gen.oper_expression

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: G-protein coupled receptor 6, Soluble cytochrome b562 chimera
hetero molecules


Theoretical massNumber of molelcules
Total (without water)50,1892
Polymers49,7151
Non-polymers4741
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)63.731, 63.731, 250.932
Angle α, β, γ (deg.)90.00, 90.00, 120.00
Int Tables number154
Space group name H-MP3221

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Components

#1: Protein G-protein coupled receptor 6, Soluble cytochrome b562 chimera / Sphingosine 1-phosphate receptor GPR6 / Cytochrome b-562


Mass: 49715.480 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human), (gene. exp.) Escherichia coli (E. coli)
Gene: GPR6, cybC / Production host: Homo sapiens (human) / References: UniProt: P46095, UniProt: P0ABE7
#2: Chemical ChemComp-X7T / 1-{2-[4-(2,4-difluorophenoxy)piperidin-1-yl]-3-{[(3R)-oxolan-3-yl]amino}-7,8-dihydropyrido[3,4-b]pyrazin-6(5H)-yl}ethan-1-one


Mass: 473.516 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C24H29F2N5O3 / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY
Has protein modificationY

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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: 2.89 Å3/Da / Density % sol: 57.43 %
Crystal growTemperature: 293 K / Method: lipidic cubic phase / Details: NaCH3COO, PEG 400, NaCl, PPG P40

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Data collection

DiffractionMean temperature: 123 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: APS / Beamline: 23-ID-B / Wavelength: 1.0332 Å
DetectorType: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Nov 13, 2020
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1.0332 Å / Relative weight: 1
ReflectionResolution: 3.49→50 Å / Num. obs: 7983 / % possible obs: 98 % / Redundancy: 5.7 % / CC1/2: 1 / Rmerge(I) obs: 0.06 / Net I/σ(I): 28.4
Reflection shellResolution: 3.5→3.56 Å / Rmerge(I) obs: 1.89 / Num. unique obs: 406 / CC1/2: 0.403

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Processing

Software
NameVersionClassification
PHENIX(1.20.1_4487: ???)refinement
PHASERphasing
HKL-2000data reduction
HKL-2000data scaling
RefinementMethod to determine structure: MOLECULAR REPLACEMENT / Resolution: 3.49→46.07 Å / SU ML: 0.68 / Cross valid method: FREE R-VALUE / σ(F): 1.33 / Phase error: 43.43 / Stereochemistry target values: ML
RfactorNum. reflection% reflection
Rfree0.3219 413 5.29 %
Rwork0.2913 --
obs0.2929 7814 96.48 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å / Solvent model: FLAT BULK SOLVENT MODEL
Refinement stepCycle: LAST / Resolution: 3.49→46.07 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms2617 0 34 0 2651
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.0042714
X-RAY DIFFRACTIONf_angle_d0.6343732
X-RAY DIFFRACTIONf_dihedral_angle_d11.643894
X-RAY DIFFRACTIONf_chiral_restr0.034467
X-RAY DIFFRACTIONf_plane_restr0.005463
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
3.49-3.560.42361320.36592377X-RAY DIFFRACTION95
3.56-5.040.37131430.31752446X-RAY DIFFRACTION98
5.04-46.070.28421380.26862578X-RAY DIFFRACTION96

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