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- PDB-6kpc: Crystal structure of an agonist bound GPCR -

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Basic information

Entry
Database: PDB / ID: 6kpc
TitleCrystal structure of an agonist bound GPCR
ComponentsCannabinoid receptor 2,Endolysin
KeywordsMEMBRANE PROTEIN / GPCR / LCP / agonist
Function / homology
Function and homology information


cannabinoid receptor activity / negative regulation of mast cell activation / negative regulation of synaptic transmission, GABAergic / negative regulation of action potential / Class A/1 (Rhodopsin-like receptors) / regulation of metabolic process / leukocyte chemotaxis / extrinsic component of cytoplasmic side of plasma membrane / G protein-coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger / viral release from host cell by cytolysis ...cannabinoid receptor activity / negative regulation of mast cell activation / negative regulation of synaptic transmission, GABAergic / negative regulation of action potential / Class A/1 (Rhodopsin-like receptors) / regulation of metabolic process / leukocyte chemotaxis / extrinsic component of cytoplasmic side of plasma membrane / G protein-coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger / viral release from host cell by cytolysis / response to amphetamine / peptidoglycan catabolic process / adenylate cyclase-activating G protein-coupled receptor signaling pathway / cell wall macromolecule catabolic process / lysozyme / lysozyme activity / G alpha (i) signalling events / perikaryon / host cell cytoplasm / response to lipopolysaccharide / defense response to bacterium / inflammatory response / immune response / dendrite / endoplasmic reticulum / plasma membrane / cytoplasm
Similarity search - Function
Cannabinoid receptor type 2 / Cannabinoid receptor family / Endolysin T4 type / T4-type lysozyme / : / Glycoside hydrolase, family 24 / Phage lysozyme / Lysozyme domain superfamily / Serpentine type 7TM GPCR chemoreceptor Srsx / G-protein coupled receptors family 1 signature. ...Cannabinoid receptor type 2 / Cannabinoid receptor family / Endolysin T4 type / T4-type lysozyme / : / Glycoside hydrolase, family 24 / Phage lysozyme / Lysozyme domain superfamily / Serpentine type 7TM GPCR chemoreceptor Srsx / 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) / Lysozyme-like domain superfamily
Similarity search - Domain/homology
Chem-E3R / Endolysin / Cannabinoid receptor 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Enterobacteria phage RB59 (virus)
MethodX-RAY DIFFRACTION / SYNCHROTRON / MOLECULAR REPLACEMENT / Resolution: 3.2 Å
AuthorsLi, X.T. / Hua, T. / Wu, L.J. / Makriyannis, A. / Wu, M. / Liu, Z.J.
CitationJournal: Cell / Year: 2020
Title: Activation and Signaling Mechanism Revealed by Cannabinoid Receptor-G Complex Structures.
Authors: Tian Hua / Xiaoting Li / Lijie Wu / Christos Iliopoulos-Tsoutsouvas / Yuxia Wang / Meng Wu / Ling Shen / Christina A Brust / Spyros P Nikas / Feng Song / Xiyong Song / Shuguang Yuan / ...Authors: Tian Hua / Xiaoting Li / Lijie Wu / Christos Iliopoulos-Tsoutsouvas / Yuxia Wang / Meng Wu / Ling Shen / Christina A Brust / Spyros P Nikas / Feng Song / Xiyong Song / Shuguang Yuan / Qianqian Sun / Yiran Wu / Shan Jiang / Travis W Grim / Othman Benchama / Edward L Stahl / Nikolai Zvonok / Suwen Zhao / Laura M Bohn / Alexandros Makriyannis / Zhi-Jie Liu /
Abstract: Human endocannabinoid systems modulate multiple physiological processes mainly through the activation of cannabinoid receptors CB1 and CB2. Their high sequence similarity, low agonist selectivity, ...Human endocannabinoid systems modulate multiple physiological processes mainly through the activation of cannabinoid receptors CB1 and CB2. Their high sequence similarity, low agonist selectivity, and lack of activation and G protein-coupling knowledge have hindered the development of therapeutic applications. Importantly, missing structural information has significantly held back the development of promising CB2-selective agonist drugs for treating inflammatory and neuropathic pain without the psychoactivity of CB1. Here, we report the cryoelectron microscopy structures of synthetic cannabinoid-bound CB2 and CB1 in complex with G, as well as agonist-bound CB2 crystal structure. Of important scientific and therapeutic benefit, our results reveal a diverse activation and signaling mechanism, the structural basis of CB2-selective agonists design, and the unexpected interaction of cholesterol with CB1, suggestive of its endogenous allosteric modulating role.
History
DepositionAug 15, 2019Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Feb 12, 2020Provider: repository / Type: Initial release
Revision 1.1Mar 11, 2020Group: Database references / Category: citation / Item: _citation.journal_volume / _citation.page_first
Revision 2.0Apr 27, 2022Group: Advisory / Atomic model ...Advisory / Atomic model / Author supporting evidence / Data collection / Database references / Derived calculations / Non-polymer description / Source and taxonomy / Structure summary
Category: atom_site / chem_comp ...atom_site / chem_comp / database_2 / entity / entity_name_com / entity_src_gen / pdbx_entity_instance_feature / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_unobs_or_zero_occ_atoms / struct_site
Item: _atom_site.auth_comp_id / _atom_site.label_comp_id ..._atom_site.auth_comp_id / _atom_site.label_comp_id / _chem_comp.formula / _chem_comp.formula_weight / _chem_comp.id / _chem_comp.mon_nstd_flag / _chem_comp.name / _chem_comp.type / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _entity.details / _entity.formula_weight / _entity.pdbx_description / _entity.pdbx_mutation / _entity_name_com.name / _entity_src_gen.gene_src_common_name / _pdbx_entity_instance_feature.auth_comp_id / _pdbx_entity_instance_feature.comp_id / _pdbx_entity_nonpoly.comp_id / _pdbx_entity_nonpoly.name / _pdbx_nonpoly_scheme.mon_id / _pdbx_nonpoly_scheme.pdb_mon_id / _struct_site.details / _struct_site.pdbx_auth_comp_id
Revision 2.1Nov 22, 2023Group: Data collection / Derived calculations ...Data collection / Derived calculations / Refinement description / Structure summary
Category: chem_comp / chem_comp_atom ...chem_comp / chem_comp_atom / chem_comp_bond / entity / pdbx_entity_nonpoly / pdbx_initial_refinement_model
Item: _chem_comp.name / _entity.pdbx_description / _pdbx_entity_nonpoly.name
Revision 2.2Oct 30, 2024Group: Structure summary / Category: pdbx_entry_details / pdbx_modification_feature / Item: _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Cannabinoid receptor 2,Endolysin
hetero molecules


Theoretical massNumber of molelcules
Total (without water)56,6102
Polymers56,2101
Non-polymers4001
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Unit cell
Length a, b, c (Å)33.960, 140.220, 156.260
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number19
Space group name H-MP212121

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Components

#1: Protein Cannabinoid receptor 2,Endolysin / hCB2 / CX5 / Lysis protein / Lysozyme / Muramidase


Mass: 56209.934 Da / Num. of mol.: 1 / Mutation: G78L,T127A,T153L,G210A,C54T,C97A,R242E,G304E
Source method: isolated from a genetically manipulated source
Details: CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 ...Details: CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion,CB2, T4 lysozyme, CB2 fusion
Source: (gene. exp.) Homo sapiens (human), (gene. exp.) Enterobacteria phage RB59 (virus)
Gene: CNR2, CB2A, CB2B, e, RB59_126 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P34972, UniProt: A0A097J809, lysozyme
#2: Chemical ChemComp-E3R / 7-[(6aR,9R,10aR)-1-Hydroxy-9-(hydroxymethyl)-6,6-dimethyl-6a,7,8,9,10,10a-hexahydro-6H-benzo[c]chromen-3-yl]- 7-methyloctanenitrile


Mass: 399.566 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C25H37NO3 / 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: 3.31 Å3/Da / Density % sol: 67.19 %
Crystal growTemperature: 293 K / Method: lipidic cubic phase
Details: 100 mM HEPES sodium pH 7.0, 25% PEG 400, 220 mM Sodium sulfate decahydrate

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: Y
Diffraction sourceSource: SYNCHROTRON / Site: SPring-8 / Beamline: BL41XU / Wavelength: 1 Å
DetectorType: DECTRIS EIGER X 16M / Detector: PIXEL / Date: Oct 19, 2018
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 1 Å / Relative weight: 1
ReflectionResolution: 3.2→44.78 Å / Num. obs: 11476 / % possible obs: 87.6 % / Redundancy: 2.847 % / Biso Wilson estimate: 96.93 Å2 / CC1/2: 0.986 / Rmerge(I) obs: 0.171 / Rrim(I) all: 0.2 / Χ2: 0.994 / Net I/σ(I): 4.26 / Num. measured all: 32673 / Scaling rejects: 21
Reflection shell

Diffraction-ID: 1

Resolution (Å)Redundancy (%)Rmerge(I) obsMean I/σ(I) obsNum. measured obsNum. possibleNum. unique obsCC1/2Rrim(I) all% possible all
3.2-3.282.7960.6731.2223859448530.6270.78990.4
3.28-3.373.0650.5991.5325999388480.6390.69790.4
3.37-3.472.70.5561.5820258857500.7350.66484.7
3.47-3.582.7780.4212.0420758457470.8990.49588.4
3.58-3.73.1320.4192.7324658737870.8480.48290.1
3.7-3.822.8670.313.2321198297390.8870.36589.1
3.82-3.972.7220.323.3318627686840.9030.3889.1
3.97-4.132.8750.2584.1920217807030.940.390.1
4.13-4.312.9110.2514.6919277386620.9410.29389.7
4.31-4.532.5230.2084.9115396906100.9370.24988.4
4.53-4.772.8440.1945.8916986935970.9470.22786.1
4.77-5.062.8510.1995.7915856305560.9640.23188.3
5.06-5.412.8760.1955.9914816095150.950.22784.6
5.41-5.842.920.1855.8814605735000.9750.21787.3
5.84-6.42.9690.1626.2913515194550.9580.18887.7
6.4-7.162.810.1546.9111694904160.9560.18384.9
7.16-8.262.8270.1458.6810464333700.9330.16985.5
8.26-10.122.6030.1228.817943683050.9570.14682.9
10.12-14.312.7470.0899.476733062450.9880.10380.1
14.31-44.782.9780.0749.883991881340.9950.08671.3
Serial crystallography sample deliveryMethod: fixed target

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Processing

Software
NameVersionClassification
BUSTER2.10.3refinement
XSCALEdata scaling
PDB_EXTRACT3.25data extraction
XDSdata reduction
PHASERphasing
RefinementMethod to determine structure: MOLECULAR REPLACEMENT
Starting model: 5zty

5zty


Resolution: 3.2→44.78 Å / Cor.coef. Fo:Fc: 0.905 / Cor.coef. Fo:Fc free: 0.871 / Cross valid method: THROUGHOUT / σ(F): 0 / SU Rfree Blow DPI: 0.48
RfactorNum. reflection% reflectionSelection details
Rfree0.262 531 4.63 %RANDOM
Rwork0.23 ---
obs0.231 11475 87.7 %-
Displacement parametersBiso max: 262.08 Å2 / Biso mean: 118.21 Å2 / Biso min: 11.04 Å2
Baniso -1Baniso -2Baniso -3
1-20.7169 Å20 Å20 Å2
2---22.8308 Å20 Å2
3---2.1138 Å2
Refine analyzeLuzzati coordinate error obs: 0.53 Å
Refinement stepCycle: final / Resolution: 3.2→44.78 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms3471 0 29 0 3500
Biso mean--114.15 --
Num. residues----445
Refine LS restraints
Refine-IDTypeNumberRestraint functionWeightDev ideal
X-RAY DIFFRACTIONt_dihedral_angle_d1211SINUSOIDAL2
X-RAY DIFFRACTIONt_trig_c_planes
X-RAY DIFFRACTIONt_gen_planes578HARMONIC5
X-RAY DIFFRACTIONt_it3581HARMONIC20
X-RAY DIFFRACTIONt_nbd
X-RAY DIFFRACTIONt_improper_torsion
X-RAY DIFFRACTIONt_pseud_angle
X-RAY DIFFRACTIONt_chiral_improper_torsion473SEMIHARMONIC5
X-RAY DIFFRACTIONt_sum_occupancies
X-RAY DIFFRACTIONt_utility_distance
X-RAY DIFFRACTIONt_utility_angle
X-RAY DIFFRACTIONt_utility_torsion
X-RAY DIFFRACTIONt_ideal_dist_contact4227SEMIHARMONIC4
X-RAY DIFFRACTIONt_bond_d3581HARMONIC20.009
X-RAY DIFFRACTIONt_angle_deg4871HARMONIC21.03
X-RAY DIFFRACTIONt_omega_torsion1.95
X-RAY DIFFRACTIONt_other_torsion19.89
LS refinement shellResolution: 3.2→3.5 Å / Rfactor Rfree error: 0 / Total num. of bins used: 6
RfactorNum. reflection% reflection
Rfree0.2744 130 4.83 %
Rwork0.2316 2560 -
all0.2337 2690 -
obs--88.31 %
Refinement TLS params.

Method: refined / 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)
14.1904-1.0658-0.37887.0911-5.820810.97730.18120.01180.15150.1045-0.5129-0.2743-0.63840.84620.3318-0.4219-0.05990.0676-0.11010.304-0.313310.6139-2.8231-32.6508
24.27570.0526-1.6961.13690.59831.75620.1452-0.54180.37250.18310.07990.1184-0.15730.0554-0.225-0.1209-0.05780.04580.0746-0.1068-0.2662-5.1493-22.92519.9196
Refinement TLS group
IDRefine-IDRefine TLS-IDSelection detailsAuth asym-IDAuth seq-ID
1X-RAY DIFFRACTION1{ A|19 - 315 }A19 - 315
2X-RAY DIFFRACTION2{ A|1001 - 1160 }A1001 - 1160

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