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- PDB-8sai: Cryo-EM structure of GPR34-Gi complex -

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Basic information

Entry
Database: PDB / ID: 8sai
TitleCryo-EM structure of GPR34-Gi complex
Components
  • (Guanine nucleotide-binding protein ...) x 3
  • Probable G-protein coupled receptor 34
  • scFv16
KeywordsLIPID BINDING PROTEIN / Complex / Agonist / MEMBRANE PROTEIN
Function / homology
Function and homology information


G protein-coupled purinergic nucleotide receptor activity / regulation of eating behavior / adenylate cyclase inhibitor activity / T cell migration / positive regulation of protein localization to cell cortex / positive regulation of relaxation of smooth muscle / Adenylate cyclase inhibitory pathway / D2 dopamine receptor binding / adenylate cyclase-inhibiting serotonin receptor signaling pathway / G protein-coupled serotonin receptor binding ...G protein-coupled purinergic nucleotide receptor activity / regulation of eating behavior / adenylate cyclase inhibitor activity / T cell migration / positive regulation of protein localization to cell cortex / positive regulation of relaxation of smooth muscle / Adenylate cyclase inhibitory pathway / D2 dopamine receptor binding / adenylate cyclase-inhibiting serotonin receptor signaling pathway / G protein-coupled serotonin receptor binding / cellular response to forskolin / regulation of mitotic spindle organization / mast cell degranulation / chemokine-mediated signaling pathway / neuropeptide signaling pathway / Regulation of insulin secretion / response to prostaglandin E / positive regulation of cholesterol biosynthetic process / G protein-coupled receptor binding / response to peptide hormone / G protein-coupled receptor activity / G-protein beta/gamma-subunit complex binding / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / Olfactory Signaling Pathway / Activation of the phototransduction cascade / G protein-coupled acetylcholine receptor signaling pathway / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / G-protein activation / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / Prostacyclin signalling through prostacyclin receptor / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / GDP binding / G beta:gamma signalling through BTK / photoreceptor disc membrane / ADP signalling through P2Y purinoceptor 12 / Glucagon-type ligand receptors / Sensory perception of sweet, bitter, and umami (glutamate) taste / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / G alpha (z) signalling events / cellular response to catecholamine stimulus / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / ADORA2B mediated anti-inflammatory cytokines production / adenylate cyclase-activating dopamine receptor signaling pathway / cellular response to prostaglandin E stimulus / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / GPER1 signaling / heterotrimeric G-protein complex / Inactivation, recovery and regulation of the phototransduction cascade / G alpha (12/13) signalling events / G-protein beta-subunit binding / extracellular vesicle / Thrombin signalling through proteinase activated receptors (PARs) / signaling receptor complex adaptor activity / adenylate cyclase-activating G protein-coupled receptor signaling pathway / GTPase binding / G protein activity / midbody / Ca2+ pathway / cell cortex / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / G alpha (i) signalling events / ciliary basal body / G alpha (s) signalling events / G alpha (q) signalling events / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / Ras protein signal transduction / Extra-nuclear estrogen signaling / G protein-coupled receptor signaling pathway / cell division / lysosomal membrane / GTPase activity / centrosome / synapse / GTP binding / protein-containing complex binding / magnesium ion binding / signal transduction / extracellular exosome / membrane / nucleus / plasma membrane / cytosol / cytoplasm
Similarity search - Function
: / G-protein alpha subunit, group I / G protein alpha subunit, helical insertion / G protein alpha subunit / Guanine nucleotide binding protein (G-protein), alpha subunit / G-protein alpha subunit / G-alpha domain profile. / G-protein, gamma subunit / G-protein gamma subunit domain profile. / G-protein gamma-like domain ...: / G-protein alpha subunit, group I / G protein alpha subunit, helical insertion / G protein alpha subunit / Guanine nucleotide binding protein (G-protein), alpha subunit / G-protein alpha subunit / G-alpha domain profile. / G-protein, gamma subunit / G-protein gamma subunit domain profile. / G-protein gamma-like domain / G-protein gamma-like domain superfamily / GGL domain / G protein gamma subunit-like motifs / GGL domain / G protein beta WD-40 repeat protein / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / G-protein coupled receptors family 1 signature. / 7 transmembrane receptor (rhodopsin family) / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Chem-S12 / 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(i) subunit alpha-1 / Probable G protein-coupled receptor 34
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.27 Å
AuthorsYong, X.H. / Zhao, C. / Yan, W. / Shao, Z.H.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC) China
CitationJournal: Proc Natl Acad Sci U S A / Year: 2023
Title: Cryo-EM structures of human GPR34 enable the identification of selective antagonists.
Authors: Anjie Xia / Xihao Yong / Changbin Zhang / Guifeng Lin / Guowen Jia / Chang Zhao / Xin Wang / Yize Hao / Yifei Wang / Pei Zhou / Xin Yang / Yue Deng / Chao Wu / Yujiao Chen / Jiawei Zhu / ...Authors: Anjie Xia / Xihao Yong / Changbin Zhang / Guifeng Lin / Guowen Jia / Chang Zhao / Xin Wang / Yize Hao / Yifei Wang / Pei Zhou / Xin Yang / Yue Deng / Chao Wu / Yujiao Chen / Jiawei Zhu / Xiaodi Tang / Jingming Liu / Shiyu Zhang / Jiahao Zhang / Zheng Xu / Qian Hu / Jinlong Zhao / Yuting Yue / Wei Yan / Zhaoming Su / Yuquan Wei / Rongbin Zhou / Haohao Dong / Zhenhua Shao / Shengyong Yang /
Abstract: GPR34 is a functional G-protein-coupled receptor of Lysophosphatidylserine (LysoPS), and has pathogenic roles in numerous diseases, yet remains poorly targeted. We herein report a cryo-electron ...GPR34 is a functional G-protein-coupled receptor of Lysophosphatidylserine (LysoPS), and has pathogenic roles in numerous diseases, yet remains poorly targeted. We herein report a cryo-electron microscopy (cryo-EM) structure of GPR34 bound with LysoPS (18:1) and G protein, revealing a unique ligand recognition mode with the negatively charged head group of LysoPS occupying a polar cavity formed by TM3, 6 and 7, and the hydrophobic tail of LysoPS residing in a lateral open hydrophobic groove formed by TM3-5. Virtual screening and subsequent structural optimization led to the identification of a highly potent and selective antagonist (YL-365). Design of fusion proteins allowed successful determination of the challenging cryo-EM structure of the inactive GPR34 complexed with YL-365, which revealed the competitive binding of YL-365 in a portion of the orthosteric binding pocket of GPR34 and the antagonist-binding-induced allostery in the receptor, implicating the inhibition mechanism of YL-365. Moreover, YL-365 displayed excellent activity in a neuropathic pain model without obvious toxicity. Collectively, this study offers mechanistic insights into the endogenous agonist recognition and antagonist inhibition of GPR34, and provides proof of concept that targeting GPR34 represents a promising strategy for disease treatment.
History
DepositionApr 1, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 4, 2023Provider: repository / Type: Initial release
Revision 1.1Nov 13, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _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: Probable G-protein coupled receptor 34
C: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
D: Guanine nucleotide-binding protein G(i) subunit alpha-1
E: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
S: scFv16
hetero molecules


Theoretical massNumber of molelcules
Total (without water)169,8956
Polymers169,3715
Non-polymers5241
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Guanine nucleotide-binding protein ... , 3 types, 3 molecules CDE

#2: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / G gamma-I


Mass: 7861.143 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
#3: Protein Guanine nucleotide-binding protein G(i) subunit alpha-1 / Adenylate cyclase-inhibiting G alpha protein


Mass: 40415.031 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNAI1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P63096
#4: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 39141.793 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

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Protein / Antibody / Non-polymers , 3 types, 3 molecules AS

#1: Protein Probable G-protein coupled receptor 34


Mass: 53316.676 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GPR34 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9UPC5
#5: Antibody scFv16


Mass: 28636.793 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Mus musculus (house mouse) / Production host: Spodoptera frugiperda (fall armyworm)
#6: Chemical ChemComp-S12 / O-[(S)-hydroxy{[(2S)-2-hydroxy-3-(octadec-9-enoyloxy)propyl]oxy}phosphoryl]-L-serine / 1-oleoyl-2-hydroxy-sn-glycero-3-phospho-L-serine


Type: L-peptide linking / Mass: 523.597 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C24H46NO9P / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY
Has protein modificationY

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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
1Human GPR34 in complex with Gi heterotrimer and scFv16COMPLEX#1-#50RECOMBINANT
2Soluble cytochrome b562,Probable G-protein coupled receptor 34, Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2, Guanine nucleotide-binding protein G(i) subunit alpha-1, Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1COMPLEX#1-#41RECOMBINANT
3scFv16COMPLEX#51RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Mus musculus (house mouse)10090
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Spodoptera frugiperda (fall armyworm)7108
23Spodoptera frugiperda (fall armyworm)7108
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 FIELD / Nominal defocus max: 1800 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 65 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.27 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 183178 / Symmetry type: POINT

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