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- PDB-8i7v: Cryo-EM structure of Acipimox bound human hydroxy-carboxylic acid... -

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Basic information

Entry
Database: PDB / ID: 8i7v
TitleCryo-EM structure of Acipimox bound human hydroxy-carboxylic acid receptor 2 in complex with Gi heterotrimer
Components
  • (Guanine nucleotide-binding protein ...) x 3
  • Human hydroxycarboxylic acid receptor 2,Hydroxycarboxylic acid receptor 2,Hydroxycarboxylic acid receptor 2
  • scFv16
KeywordsMEMBRANE PROTEIN / GPCR / G-Protein / acipimox / signaling
Function / homology
Function and homology information


nicotinic acid receptor activity / Hydroxycarboxylic acid-binding receptors / neutrophil apoptotic process / Class A/1 (Rhodopsin-like receptors) / positive regulation of neutrophil apoptotic process / positive regulation of adiponectin secretion / Adenylate cyclase inhibitory pathway / positive regulation of protein localization to cell cortex / regulation of cAMP-mediated signaling / D2 dopamine receptor binding ...nicotinic acid receptor activity / Hydroxycarboxylic acid-binding receptors / neutrophil apoptotic process / Class A/1 (Rhodopsin-like receptors) / positive regulation of neutrophil apoptotic process / positive regulation of adiponectin secretion / Adenylate cyclase inhibitory pathway / positive regulation of protein localization to cell cortex / regulation of cAMP-mediated signaling / D2 dopamine receptor binding / G protein-coupled serotonin receptor binding / negative regulation of lipid catabolic process / regulation of mitotic spindle organization / cellular response to forskolin / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / Regulation of insulin secretion / G protein-coupled receptor binding / 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 / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / 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 / ADP signalling through P2Y purinoceptor 12 / G beta:gamma signalling through BTK / Sensory perception of sweet, bitter, and umami (glutamate) taste / response to peptide hormone / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / photoreceptor disc membrane / Adrenaline,noradrenaline inhibits insulin secretion / Glucagon-type ligand receptors / Vasopressin regulates renal water homeostasis via Aquaporins / 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 / GDP binding / 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 / Thrombin signalling through proteinase activated receptors (PARs) / retina development in camera-type eye / GTPase binding / cell junction / Ca2+ pathway / phospholipase C-activating G protein-coupled receptor signaling pathway / cell cortex / midbody / 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 / cell cycle / G protein-coupled receptor signaling pathway / cell division / lysosomal membrane / GTPase activity / centrosome / synapse / protein-containing complex binding / GTP binding / nucleolus / magnesium ion binding / signal transduction / extracellular exosome / nucleoplasm / membrane / plasma membrane / cytosol / cytoplasm
Similarity search - Function
G-protein alpha subunit, group I / 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 alpha subunit, group I / 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) / 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
Similarity search - Domain/homology
Chem-OJX / 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 / Hydroxycarboxylic acid receptor 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.77 Å
AuthorsPark, J.H. / Ishimoto, N. / Park, S.Y.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nat Commun / Year: 2023
Title: Structural basis for ligand recognition and signaling of hydroxy-carboxylic acid receptor 2.
Authors: Jae-Hyun Park / Kouki Kawakami / Naito Ishimoto / Tatsuya Ikuta / Mio Ohki / Toru Ekimoto / Mitsunori Ikeguchi / Dong-Sun Lee / Young-Ho Lee / Jeremy R H Tame / Asuka Inoue / Sam-Yong Park /
Abstract: Hydroxycarboxylic acid receptors (HCAR1, HCAR2, and HCAR3) transduce G signaling upon biding to molecules such as lactic acid, butyric acid and 3-hydroxyoctanoic acid, which are associated with ...Hydroxycarboxylic acid receptors (HCAR1, HCAR2, and HCAR3) transduce G signaling upon biding to molecules such as lactic acid, butyric acid and 3-hydroxyoctanoic acid, which are associated with lipolytic and atherogenic activity, and neuroinflammation. Although many reports have elucidated the function of HCAR2 and its potential as a therapeutic target for treating not only dyslipidemia but also neuroimmune disorders such as multiple sclerosis and Parkinson's disease, the structural basis of ligand recognition and ligand-induced G-coupling remains unclear. Here we report three cryo-EM structures of the human HCAR2-G signaling complex, each bound with different ligands: niacin, acipimox or GSK256073. All three agonists are held in a deep pocket lined by residues that are not conserved in HCAR1 and HCAR3. A distinct hairpin loop at the HCAR2 N-terminus and extra-cellular loop 2 (ECL2) completely enclose the ligand. These structures also reveal the agonist-induced conformational changes propagated to the G-protein-coupling interface during activation. Collectively, the structures presented here are expected to help in the design of ligands specific for HCAR2, leading to new drugs for the treatment of various diseases such as dyslipidemia and inflammation.
History
DepositionFeb 2, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Feb 7, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Human hydroxycarboxylic acid receptor 2,Hydroxycarboxylic acid receptor 2,Hydroxycarboxylic acid receptor 2
B: Guanine nucleotide-binding protein G(i) subunit alpha-1
C: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
D: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
E: scFv16
hetero molecules


Theoretical massNumber of molelcules
Total (without water)168,5356
Polymers168,3805
Non-polymers1551
Water0
1


  • Idetical with deposited unit
  • defined by author&software
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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

#2: 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
#3: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 37728.152 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: 8506.765 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

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

#1: Protein Human hydroxycarboxylic acid receptor 2,Hydroxycarboxylic acid receptor 2,Hydroxycarboxylic acid receptor 2 / G-protein coupled receptor 109A / G-protein coupled receptor HM74A / Niacin receptor 1 / Nicotinic ...G-protein coupled receptor 109A / G-protein coupled receptor HM74A / Niacin receptor 1 / Nicotinic acid receptor


Mass: 54320.297 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: HCAR2, GPR109A, HCA2, HM74A, NIACR1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q8TDS4
#5: Antibody scFv16


Mass: 27409.588 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-OJX / 5-methyl-4-oxidanyl-pyrazin-4-ium-2-carboxylic acid


Mass: 155.131 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C6H7N2O3

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Details

Has ligand of interestY

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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

ComponentName: Acipomox bound human hydroxy-carboxylic acid receptor 2 in complex with Gi heterotrimer
Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT
Molecular weightValue: 0.15 MDa / Experimental value: YES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
Details: 20 mM HEPES pH8.0, 100 mM NaCl, 1 mM MgCl2, 0.5 mM TCEP, 0.001% LMNG, 0.0001% CHS
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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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 / Nominal magnification: 105000 X / Nominal defocus max: 1800 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameCategory
1cryoSPARCparticle selection
2EPUimage acquisition
4cryoSPARCCTF correction
7UCSF ChimeraXmodel fitting
8Cootmodel fitting
10PHENIXmodel refinement
11cryoSPARCinitial Euler assignment
12cryoSPARCfinal Euler assignment
13RELIONclassification
14cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.77 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 228127 / Num. of class averages: 2 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0059244
ELECTRON MICROSCOPYf_angle_d0.65612505
ELECTRON MICROSCOPYf_dihedral_angle_d5.1491236
ELECTRON MICROSCOPYf_chiral_restr0.0451401
ELECTRON MICROSCOPYf_plane_restr0.0051582

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