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- EMDB-39281: A3R-Gi complex bound to i6A -

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Basic information

Entry
Database: EMDB / ID: EMD-39281
TitleA3R-Gi complex bound to i6A
Map data
Sample
  • Complex: A3R-Gi complex bound to i6a
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2,Guanine nucleotide-binding protein G(i) subunit alpha-1 chimera
    • Protein or peptide: Hemagglutinin,Adenosine receptor A3,GFP chimera
    • Protein or peptide: scfv16
  • Ligand: N-(3-methylbut-2-en-1-yl)adenosine
KeywordsGPCR / G protein / adenosine receptor / adenosine / complex / SIGNALING PROTEIN
Function / homology
Function and homology information


Adenylate cyclase inhibitory pathway / Olfactory Signaling Pathway / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / G protein-coupled adenosine receptor activity / Activation of the phototransduction cascade / Activation of G protein gated Potassium channels / G-protein activation / G beta:gamma signalling through PI3Kgamma / Prostacyclin signalling through prostacyclin receptor ...Adenylate cyclase inhibitory pathway / Olfactory Signaling Pathway / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / G protein-coupled adenosine receptor activity / Activation of the phototransduction cascade / Activation of G protein gated Potassium channels / G-protein activation / G beta:gamma signalling through PI3Kgamma / Prostacyclin signalling through prostacyclin receptor / G beta:gamma signalling through PLC beta / ADP signalling through P2Y purinoceptor 1 / Thromboxane signalling through TP receptor / Presynaptic function of Kainate receptors / G beta:gamma signalling through CDC42 / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / G alpha (12/13) signalling events / Glucagon-type ligand receptors / G beta:gamma signalling through BTK / ADP signalling through P2Y purinoceptor 12 / Adrenaline,noradrenaline inhibits insulin secretion / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / Ca2+ pathway / Thrombin signalling through proteinase activated receptors (PARs) / G alpha (z) signalling events / Extra-nuclear estrogen signaling / G alpha (s) signalling events / G alpha (q) signalling events / G alpha (i) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / Vasopressin regulates renal water homeostasis via Aquaporins / positive regulation of protein localization to cell cortex / G protein-coupled serotonin receptor binding / cellular response to forskolin / regulation of mitotic spindle organization / viral budding from plasma membrane / bioluminescence / generation of precursor metabolites and energy / G protein-coupled receptor binding / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G-protein beta/gamma-subunit complex binding / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / G-protein activation / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / G beta:gamma signalling through CDC42 / Glucagon signaling in metabolic regulation / G beta:gamma signalling through BTK / ADP signalling through P2Y purinoceptor 12 / photoreceptor disc membrane / Glucagon-type ligand receptors / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / GDP binding / G alpha (z) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / cellular response to catecholamine stimulus / ADORA2B mediated anti-inflammatory cytokines production / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / adenylate cyclase-activating dopamine receptor signaling pathway / GPER1 signaling / cellular response to prostaglandin E stimulus / G-protein beta-subunit binding / heterotrimeric G-protein complex / G alpha (12/13) signalling events / sensory perception of taste / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / GTPase binding / Ca2+ pathway / retina development in camera-type eye / midbody / cell cortex / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / fibroblast proliferation / G alpha (i) signalling events / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / G alpha (s) signalling events / phospholipase C-activating G protein-coupled receptor signaling pathway / G alpha (q) signalling events / clathrin-dependent endocytosis of virus by host cell / Extra-nuclear estrogen signaling / cell population proliferation / host cell surface receptor binding / G protein-coupled receptor signaling pathway / cell division / fusion of virus membrane with host plasma membrane / GTPase activity / fusion of virus membrane with host endosome membrane / synapse / viral envelope / centrosome / dendrite / protein-containing complex binding
Similarity search - Function
Adenosine A3 receptor / Adenosine receptor / Haemagglutinin, influenzavirus A / Haemagglutinin, HA1 chain, alpha/beta domain superfamily / Haemagglutinin / Haemagglutinin, influenzavirus A/B / Viral capsid/haemagglutinin protein / Green fluorescent protein, GFP / G-protein alpha subunit, group I / Green fluorescent protein-related ...Adenosine A3 receptor / Adenosine receptor / Haemagglutinin, influenzavirus A / Haemagglutinin, HA1 chain, alpha/beta domain superfamily / Haemagglutinin / Haemagglutinin, influenzavirus A/B / Viral capsid/haemagglutinin protein / Green fluorescent protein, GFP / G-protein alpha subunit, group I / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Serpentine type 7TM GPCR chemoreceptor Srsx / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / 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 / 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 / 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
GFP / Hemagglutinin / 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 / Adenosine receptor A3
Similarity search - Component
Biological speciesOvis aries (sheep) / Rattus rattus (black rat) / Bos taurus (domestic cattle) / Mus musculus (house mouse) / human respiratory syncytial virus
Methodsingle particle reconstruction / cryo EM / Resolution: 3.66 Å
AuthorsOshima HS / Shihoya W / Nureki O
Funding support Japan, 1 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)21H05037 Japan
CitationJournal: Nat Commun / Year: 2024
Title: Structural insights into the agonist selectivity of the adenosine A receptor.
Authors: Hidetaka S Oshima / Akiko Ogawa / Fumiya K Sano / Hiroaki Akasaka / Tomoyoshi Kawakami / Aika Iwama / Hiroyuki H Okamoto / Chisae Nagiri / Fan-Yan Wei / Wataru Shihoya / Osamu Nureki /
Abstract: Adenosine receptors play pivotal roles in physiological processes. Adenosine A receptor (AR), the most recently identified adenosine receptor, is expressed in various tissues, exhibiting important ...Adenosine receptors play pivotal roles in physiological processes. Adenosine A receptor (AR), the most recently identified adenosine receptor, is expressed in various tissues, exhibiting important roles in neuron, heart, and immune cells, and is often overexpressed in tumors, highlighting the therapeutic potential of AR-selective agents. Recently, we identified RNA-derived N-methyladenosine (mA) as an endogenous agonist for AR, suggesting the relationship between RNA-derived modified adenosine and AR. Despite extensive studies on the other adenosine receptors, the selectivity mechanism of AR, especially for AR-selective agonists such as mA and namodenoson, remained elusive. Here, we identify tRNA-derived N-isopentenyl adenosine (iA) as an AR-selective ligand via screening of modified nucleosides against the adenosine receptors. Like mA, iA is found in the human body and may be an endogenous AR ligand. Our cryo-EM analyses elucidate the AR-G complexes bound to adenosine, 5'-N-ethylcarboxamidoadenosine (NECA), mA, iA, and namodenoson at overall resolutions of 3.27 Å (adenosine), 2.86 Å (NECA), 3.19 Å (mA), 3.28 Å (iA), and 3.20 Å (namodenoson), suggesting the selectivity and activation mechanism of AR. We further conduct structure-guided engineering of mA-insensitive AR, which may aid future research targeting mA and AR, providing a molecular basis for future drug discovery.
History
DepositionFeb 27, 2024-
Header (metadata) releaseNov 6, 2024-
Map releaseNov 6, 2024-
UpdateJun 18, 2025-
Current statusJun 18, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_39281.map.gz / Format: CCP4 / Size: 35.3 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
1.11 Å/pix.
x 210 pix.
= 233.1 Å
1.11 Å/pix.
x 210 pix.
= 233.1 Å
1.11 Å/pix.
x 210 pix.
= 233.1 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.11 Å
Density
Contour LevelBy AUTHOR: 5.07
Minimum - Maximum-21.10474 - 37.661625000000001
Average (Standard dev.)0.0019296176 (±1.084067)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin000
Dimensions210210210
Spacing210210210
CellA=B=C: 233.1 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : A3R-Gi complex bound to i6a

EntireName: A3R-Gi complex bound to i6a
Components
  • Complex: A3R-Gi complex bound to i6a
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
    • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2,Guanine nucleotide-binding protein G(i) subunit alpha-1 chimera
    • Protein or peptide: Hemagglutinin,Adenosine receptor A3,GFP chimera
    • Protein or peptide: scfv16
  • Ligand: N-(3-methylbut-2-en-1-yl)adenosine

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Supramolecule #1: A3R-Gi complex bound to i6a

SupramoleculeName: A3R-Gi complex bound to i6a / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2, #4, #3
Source (natural)Organism: Ovis aries (sheep)

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Macromolecule #1: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1

MacromoleculeName: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Rattus rattus (black rat)
Molecular weightTheoretical: 40.586332 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGSLLQSELD QLRQEAEQLK NQIRDARKAC ADATLSQITN NIDPVGRIQM RTRRTLRGHL AKIYAMHWGT DSRLLVSASQ DGKLIIWDS YTTNKVHAIP LRSSWVMTCA YAPSGNYVAC GGLDNICSIY NLKTREGNVR VSRELAGHTG YLSCCRFLDD N QIVTSSGD ...String:
MGSLLQSELD QLRQEAEQLK NQIRDARKAC ADATLSQITN NIDPVGRIQM RTRRTLRGHL AKIYAMHWGT DSRLLVSASQ DGKLIIWDS YTTNKVHAIP LRSSWVMTCA YAPSGNYVAC GGLDNICSIY NLKTREGNVR VSRELAGHTG YLSCCRFLDD N QIVTSSGD TTCALWDIET GQQTTTFTGH TGDVMSLSLA PDTRLFVSGA CDASAKLWDV REGMCRQTFT GHESDINAIC FF PNGNAFA TGSDDATCRL FDLRADQELM TYSHDNIICG ITSVSFSKSG RLLLAGYDDF NCNVWDALKA DRAGVLAGHD NRV SCLGVT DDGMAVATGS WDSFLKIWNG ASGGGSGGNS GSSGGSSGVS GWRLFKKIS

UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1

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

MacromoleculeName: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2,Guanine nucleotide-binding protein G(i) subunit alpha-1 chimera
type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Bos taurus (domestic cattle)
Molecular weightTheoretical: 48.846695 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MASNNTASIA QARKLVEQLK MEANIDRIKV SKAAADLMAY CEAHAKEDPL LTPVPASENP FREKKFFCAI LGSAGSAGSA MCTLSAEDK AAVERSKMID RNLREDGEKA AREVKLLLLG AGESGKSTIV KQMKIIHEAG YSEEECKQYK AVVYSNTIQS I IAIIRAMG ...String:
MASNNTASIA QARKLVEQLK MEANIDRIKV SKAAADLMAY CEAHAKEDPL LTPVPASENP FREKKFFCAI LGSAGSAGSA MCTLSAEDK AAVERSKMID RNLREDGEKA AREVKLLLLG AGESGKSTIV KQMKIIHEAG YSEEECKQYK AVVYSNTIQS I IAIIRAMG RLKIDFGDSA RADDARQLFV LAGAAEEGFM TAELAGVIKR LWKDSGVQAC FNRSREYQLN DSAAYYLNDL DR IAQPNYI PTQQDVLRTR VKTTGIVETH FTFKDLHFKM FDVGGQRSER KKWIHCFEGV TAIIFCVALS DYDLVLAEDE EMN RMHESM KLFDSICNNK WFTDTSIILF LNKKDLFEEK IKKSPLTICY PEYAGSNTYE EAAAYIQCQF EDLNKRKDTK EIYT HFTCA TDTKNVQFVF DAVTDVIIKN NLKDCGLF

UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2, Guanine nucleotide-binding protein G(i) subunit alpha-1

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Macromolecule #3: scfv16

MacromoleculeName: scfv16 / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 27.720795 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DVQLVESGGG LVQPGGSRKL SCSASGFAFS SFGMHWVRQA PEKGLEWVAY ISSGSGTIYY ADTVKGRFTI SRDDPKNTLF LQMTSLRSE DTAMYYCVRS IYYYGSSPFD FWGQGTTLTV SSGGGGSGGG GSGGGGSDIV MTQATSSVPV TPGESVSISC R SSKSLLHS ...String:
DVQLVESGGG LVQPGGSRKL SCSASGFAFS SFGMHWVRQA PEKGLEWVAY ISSGSGTIYY ADTVKGRFTI SRDDPKNTLF LQMTSLRSE DTAMYYCVRS IYYYGSSPFD FWGQGTTLTV SSGGGGSGGG GSGGGGSDIV MTQATSSVPV TPGESVSISC R SSKSLLHS NGNTYLYWFL QRPGQSPQLL IYRMSNLASG VPDRFSGSGS GTAFTLTISR LEAEDVGVYY CMQHLEYPLT FG AGTKLEL KAAAASSEDL YFQ

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Macromolecule #4: Hemagglutinin,Adenosine receptor A3,GFP chimera

MacromoleculeName: Hemagglutinin,Adenosine receptor A3,GFP chimera / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: human respiratory syncytial virus
Molecular weightTheoretical: 88.315891 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MKTIIALSYI FCLVFADYKD DDDAMGPVNS TAVSWTSVTY ITVEILIGLC AIVGNVLVIW VVKLNPSLQT TTFYFIVSLA LADIAVGVL VMPLAIVISL GVTIHFYSCL FMTCLMLIFT HASIMSLLAI AVDRYLRVKL TVRYRRVTTQ RRIWLALGLC W LVSFLVGL ...String:
MKTIIALSYI FCLVFADYKD DDDAMGPVNS TAVSWTSVTY ITVEILIGLC AIVGNVLVIW VVKLNPSLQT TTFYFIVSLA LADIAVGVL VMPLAIVISL GVTIHFYSCL FMTCLMLIFT HASIMSLLAI AVDRYLRVKL TVRYRRVTTQ RRIWLALGLC W LVSFLVGL TPMFGWNMKL SSADENLTFL PCRFRSVMRM DYMVYFSFFL WILVPLVVMC AIYFDIFYII RNRLSQSFSG SR ETGAFYG REFKTAKSLL LVLFLFALCW LPLSIINCIL YFDGQVPQTV LYLGILLSHA NSMMNPIVYA YKIKKFKETY LLI LKACVI CQPSKSMDPS TEQTSEGSGG GGSGGSSSGG VFTLEDFVGD WEQTAAYNLD QVLEQGGVSS LLQNLAVSVT PIQR IVRSG ENALKIDIHV IIPYEGLSAD QMAQIEEVFK VVYPVDDHHF KVILPYGTLV IDGVTPNMLN YFGRPYEGIA VFDGK KITV TGTLWNGNKI IDERLITPDG SMLFRVTINS GGSGGGGSGG SSSGGLEVLF QGPGSAAAAV SKGEELFTGV VPILVE LDG DVNGHKFSVS GEGEGDATYG KLTLKFICTT GKLPVPWPTL VTTLTYGVQC FSRYPDHMKQ HDFFKSAMPE GYVQERT IF FKDDGNYKTR AEVKFEGDTL VNRIELKGID FKEDGNILGH KLEYNYNSHN VYIMADKQKN GIKVNFKIRH NIEDGSVQ L ADHYQQNTPI GDGPVLLPDN HYLSTQSKLS KDPNEKRDHM VLLEFVTAAG ITLGMDELYK SGLRSHHHHH HHH

UniProtKB: Hemagglutinin, Adenosine receptor A3, GFP

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Macromolecule #5: N-(3-methylbut-2-en-1-yl)adenosine

MacromoleculeName: N-(3-methylbut-2-en-1-yl)adenosine / type: ligand / ID: 5 / Number of copies: 1 / Formula: ZIR
Molecular weightTheoretical: 335.358 Da
Chemical component information

ChemComp-ZIR:
N-(3-methylbut-2-en-1-yl)adenosine

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.6 µm / Nominal defocus min: 0.8 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.66 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 141536
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD

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