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- PDB-6li3: cryo-EM structure of GPR52-miniGs-NB35 -

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Basic information

Entry
Database: PDB / ID: 6li3
Titlecryo-EM structure of GPR52-miniGs-NB35
Components
  • G-protein coupled receptor 52G protein-coupled receptor
  • Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
  • Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
  • Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
  • nanobody Nb35
KeywordsMEMBRANE PROTEIN / G-protein coupled receptor / orphan GPCR / self-activation / cryo-EM
Function / homology
Function and homology information


G protein-coupled photoreceptor activity / cellular response to light stimulus / PKA activation in glucagon signalling / phototransduction / hair follicle placode formation / intracellular transport / D1 dopamine receptor binding / developmental growth / Hedgehog 'off' state / positive regulation of cAMP-mediated signaling ...G protein-coupled photoreceptor activity / cellular response to light stimulus / PKA activation in glucagon signalling / phototransduction / hair follicle placode formation / intracellular transport / D1 dopamine receptor binding / developmental growth / Hedgehog 'off' state / positive regulation of cAMP-mediated signaling / adenylate cyclase-activating adrenergic receptor signaling pathway / activation of adenylate cyclase activity / adenylate cyclase activator activity / trans-Golgi network membrane / locomotory behavior / G protein-coupled receptor activity / G-protein beta/gamma-subunit complex binding / Olfactory Signaling Pathway / Activation of the phototransduction cascade / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / bone development / G-protein activation / G protein-coupled acetylcholine receptor signaling pathway / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / adenylate cyclase-activating G protein-coupled receptor signaling pathway / ADP signalling through P2Y purinoceptor 12 / G beta:gamma signalling through BTK / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / photoreceptor disc membrane / Adrenaline,noradrenaline inhibits insulin secretion / platelet aggregation / Glucagon-type ligand receptors / cognition / Vasopressin regulates renal water homeostasis via Aquaporins / positive regulation of GTPase activity / G alpha (z) signalling events / cellular response to catecholamine stimulus / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / ADORA2B mediated anti-inflammatory cytokines production / adenylate cyclase-activating dopamine receptor signaling pathway / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / cellular response to prostaglandin E stimulus / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / sensory perception of taste / GPER1 signaling / G-protein beta-subunit binding / heterotrimeric G-protein complex / Inactivation, recovery and regulation of the phototransduction cascade / extracellular vesicle / G alpha (12/13) signalling events / signaling receptor complex adaptor activity / sensory perception of smell / Thrombin signalling through proteinase activated receptors (PARs) / retina development in camera-type eye / GTPase binding / Ca2+ pathway / phospholipase C-activating G protein-coupled receptor signaling pathway / positive regulation of cold-induced thermogenesis / G alpha (i) signalling events / fibroblast proliferation / G alpha (s) signalling events / G alpha (q) signalling events / Ras protein signal transduction / cell population proliferation / Extra-nuclear estrogen signaling / response to xenobiotic stimulus / G protein-coupled receptor signaling pathway / lysosomal membrane / GTPase activity / synapse / protein-containing complex binding / GTP binding / signal transduction / extracellular exosome / membrane / metal ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Transducin (heterotrimeric G protein), gamma chain / G Protein Gi Gamma 2 / G-protein alpha subunit, group S / YVTN repeat-like/Quinoprotein amine dehydrogenase / 7 Propeller / Methylamine Dehydrogenase; Chain H / G-alpha domain profile. / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit ...Transducin (heterotrimeric G protein), gamma chain / G Protein Gi Gamma 2 / G-protein alpha subunit, group S / YVTN repeat-like/Quinoprotein amine dehydrogenase / 7 Propeller / Methylamine Dehydrogenase; Chain H / G-alpha domain profile. / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / GGL domain / G-protein gamma-like domain superfamily / G-protein gamma-like domain / GGL domain / G protein gamma subunit-like motifs / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / 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) / Few Secondary Structures / Irregular / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase / Mainly Beta
Similarity search - Domain/homology
Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Guanine nucleotide-binding protein G(s) subunit alpha isoforms short / G-protein coupled receptor 52
Similarity search - Component
Biological speciesHomo sapiens (human)
Lama glama (llama)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.32 Å
AuthorsLi, M. / Wang, N. / Xu, F. / Wu, J. / Lei, M.
CitationJournal: Nature / Year: 2020
Title: Structural basis of ligand recognition and self-activation of orphan GPR52.
Authors: Xi Lin / Mingyue Li / Niandong Wang / Yiran Wu / Zhipu Luo / Shimeng Guo / Gye-Won Han / Shaobai Li / Yang Yue / Xiaohu Wei / Xin Xie / Yong Chen / Suwen Zhao / Jian Wu / Ming Lei / Fei Xu /
Abstract: GPR52 is a class-A orphan G-protein-coupled receptor that is highly expressed in the brain and represents a promising therapeutic target for the treatment of Huntington's disease and several ...GPR52 is a class-A orphan G-protein-coupled receptor that is highly expressed in the brain and represents a promising therapeutic target for the treatment of Huntington's disease and several psychiatric disorders. Pathological malfunction of GPR52 signalling occurs primarily through the heterotrimeric G protein, but it is unclear how GPR52 and G couple for signal transduction and whether a native ligand or other activating input is required. Here we present the high-resolution structures of human GPR52 in three states: a ligand-free state, a G-coupled self-activation state and a potential allosteric ligand-bound state. Together, our structures reveal that extracellular loop 2 occupies the orthosteric binding pocket and operates as a built-in agonist, conferring an intrinsically high level of basal activity to GPR52. A fully active state is achieved when G is coupled to GPR52 in the absence of an external agonist. The receptor also features a side pocket for ligand binding. These insights into the structure and function of GPR52 could improve our understanding of other self-activated GPCRs, enable the identification of endogenous and tool ligands, and guide drug discovery efforts that target GPR52.
History
DepositionDec 10, 2019Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Feb 26, 2020Provider: repository / Type: Initial release
Revision 1.1Mar 4, 2020Group: Database references / Category: citation / citation_author
Item: _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name
Revision 1.2Mar 18, 2020Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

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Assembly

Deposited unit
R: G-protein coupled receptor 52
A: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
B: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
G: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
N: nanobody Nb35


Theoretical massNumber of molelcules
Total (without water)129,3535
Polymers129,3535
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area12470 Å2
ΔGint-60 kcal/mol
Surface area49780 Å2

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Components

#1: Protein G-protein coupled receptor 52 / G protein-coupled receptor


Mass: 39128.777 Da / Num. of mol.: 1 / Mutation: A130W, C314P
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GPR52 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9Y2T5
#2: Protein Guanine nucleotide-binding protein G(s) subunit alpha isoforms short / Adenylate cyclase-stimulating G alpha protein


Mass: 28907.684 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNAS, GNAS1, GSP / Plasmid: pET15b / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3) / References: UniProt: P63092
#3: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 37416.930 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P62873
#4: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / G gamma-I


Mass: 7845.078 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNG2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P59768
#5: Antibody nanobody Nb35


Mass: 16054.232 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Plasmid: pET26b / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): BL21(DE3)

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1GPR52-miniGs-NB35COMPLEXall0MULTIPLE SOURCES
2GPR52-GsBeta-GsGammaCOMPLEX#1, #3-#41RECOMBINANT
3GsAlpha-NB35COMPLEX#2, #51RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Spodoptera frugiperda (fall armyworm)7108
23Escherichia coli BL21(DE3) (bacteria)469008
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 40 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.32 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 651465 / Symmetry type: POINT

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