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- PDB-8uhb: Cryo-EM Structure of the Ro5256390-bound hTA1-Gs heterotrimer sig... -

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Basic information

Entry
Database: PDB / ID: 8uhb
TitleCryo-EM Structure of the Ro5256390-bound hTA1-Gs heterotrimer signaling complex
Components
  • (Guanine nucleotide-binding protein ...) x 3
  • Nanobody 35
  • Soluble cytochrome b562,Beta-2 adrenergic receptor,Trace amine-associated receptor 1
KeywordsMEMBRANE PROTEIN / GPCR / Signaling Complex / Trace Amine-associated Receptor
Function / homology
Function and homology information


Amine ligand-binding receptors / trace-amine receptor activity / beta2-adrenergic receptor activity / positive regulation of mini excitatory postsynaptic potential / AMPA selective glutamate receptor signaling pathway / adenylate cyclase-inhibiting adrenergic receptor signaling pathway / positive regulation of autophagosome maturation / norepinephrine-epinephrine-mediated vasodilation involved in regulation of systemic arterial blood pressure / norepinephrine binding / Adrenoceptors ...Amine ligand-binding receptors / trace-amine receptor activity / beta2-adrenergic receptor activity / positive regulation of mini excitatory postsynaptic potential / AMPA selective glutamate receptor signaling pathway / adenylate cyclase-inhibiting adrenergic receptor signaling pathway / positive regulation of autophagosome maturation / norepinephrine-epinephrine-mediated vasodilation involved in regulation of systemic arterial blood pressure / norepinephrine binding / Adrenoceptors / positive regulation of lipophagy / positive regulation of cardiac muscle cell contraction / negative regulation of G protein-coupled receptor signaling pathway / regulation of smooth muscle contraction / adrenergic receptor signaling pathway / response to psychosocial stress / endosome to lysosome transport / positive regulation of glutamate receptor signaling pathway / alkylglycerophosphoethanolamine phosphodiesterase activity / negative regulation of cardiac muscle cell apoptotic process / adenylate cyclase binding / potassium channel regulator activity / neuronal dense core vesicle / adenylate cyclase-activating adrenergic receptor signaling pathway / positive regulation of cardiac muscle cell apoptotic process / endomembrane system / receptor-mediated endocytosis / adenylate cyclase activator activity / electron transport chain / clathrin-coated endocytic vesicle membrane / cellular response to amyloid-beta / 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 / phospholipase C-activating 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) / 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 / Cargo recognition for clathrin-mediated endocytosis / 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 / positive regulation of cold-induced thermogenesis / amyloid-beta binding / 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 / Clathrin-mediated endocytosis / 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 / cytoplasmic vesicle / Ca2+ pathway / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / G alpha (i) signalling events / G alpha (s) signalling events / G alpha (q) signalling events / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / positive regulation of MAPK cascade / Ras protein signal transduction / electron transfer activity / periplasmic space / signaling receptor complex / cell surface receptor signaling pathway / lysosome / Extra-nuclear estrogen signaling / endosome / apical plasma membrane / endosome membrane / Ub-specific processing proteases / iron ion binding / G protein-coupled receptor signaling pathway
Similarity search - Function
Trace amine associated receptor 1 / Trace amine associated receptor family / : / Beta 2 adrenoceptor / Adrenoceptor family / G-protein alpha subunit, group S / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / Serpentine type 7TM GPCR chemoreceptor Srsx ...Trace amine associated receptor 1 / Trace amine associated receptor family / : / Beta 2 adrenoceptor / Adrenoceptor family / G-protein alpha subunit, group S / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / Serpentine type 7TM GPCR chemoreceptor Srsx / 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-WV8 / Beta-2 adrenergic receptor / Soluble cytochrome b562 / 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 / Trace amine-associated receptor 1
Similarity search - Component
Biological speciesHomo sapiens (human)
Rattus norvegicus (Norway rat)
Lama glama (llama)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.35 Å
AuthorsZilberg, G. / Warren, A.L. / Parpounas, A.K. / Wacker, D.
Funding support United States, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM133504 United States
National Institutes of Health/National Institute of Mental Health (NIH/NIMH)MH132317 United States
National Institutes of Health/National Institute on Drug Abuse (NIH/NIDA)DA053558 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM062754 United States
CitationJournal: Nat Commun / Year: 2024
Title: Molecular basis of human trace amine-associated receptor 1 activation.
Authors: Gregory Zilberg / Alexandra K Parpounas / Audrey L Warren / Shifan Yang / Daniel Wacker /
Abstract: The human trace amine-associated receptor 1 (hTAAR1, hTA1) is a key regulator of monoaminergic neurotransmission and the actions of psychostimulants. Despite preclinical research demonstrating its ...The human trace amine-associated receptor 1 (hTAAR1, hTA1) is a key regulator of monoaminergic neurotransmission and the actions of psychostimulants. Despite preclinical research demonstrating its tractability as a drug target, its molecular mechanisms of activation remain unclear. Moreover, poorly understood pharmacological differences between rodent and human TA1 complicate the translation of findings from preclinical disease models into novel pharmacotherapies. To elucidate hTA1's mechanisms on the molecular scale and investigate the underpinnings of its divergent pharmacology from rodent orthologs, we herein report the structure of the human TA1 receptor in complex with a Gαs heterotrimer. Our structure reveals shared structural elements with other TAARs, as well as with its closest monoaminergic orthologue, the serotonin receptor 5-HT4R. We further find that a single mutation dramatically shifts the selectivity of hTA1 towards that of its rodent orthologues, and report on the effects of substituting residues to those found in serotonin and dopamine receptors. Strikingly, we also discover that the atypical antipsychotic medication and pan-monoaminergic antagonist asenapine potently and efficaciously activates hTA1. Together our studies provide detailed insight into hTA1 structure and function, contrast its molecular pharmacology with that of related receptors, and uncover off-target activities of monoaminergic drugs at hTA1.
History
DepositionOct 8, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 10, 2024Provider: repository / Type: Initial release
Revision 1.1May 1, 2024Group: Database references / Category: citation
Item: _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Nov 6, 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: Soluble cytochrome b562,Beta-2 adrenergic receptor,Trace amine-associated receptor 1
B: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
C: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
G: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
N: Nanobody 35
hetero molecules


Theoretical massNumber of molelcules
Total (without water)168,0356
Polymers167,8175
Non-polymers2181
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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

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


Mass: 39418.086 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P62873
#3: Protein Guanine nucleotide-binding protein G(s) subunit alpha isoforms short


Mass: 45803.598 Da / Num. of mol.: 1
Mutation: E189D, M191V, T193S, E194D, N271K, K274D, R280K, T284D, I285T, G226A, A366S
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Gnas / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P63095
#4: 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

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

#1: Protein Soluble cytochrome b562,Beta-2 adrenergic receptor,Trace amine-associated receptor 1 / Cytochrome b-562


Mass: 60019.816 Da / Num. of mol.: 1 / Mutation: M(-129)W, R(-14)G, Q(-3)E, F112W
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: cybC, ADRB2, TAAR1 / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: P0ABE7, UniProt: P07550, UniProt: Q96RJ0
#5: Antibody Nanobody 35


Mass: 14714.320 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Production host: Spodoptera frugiperda (fall armyworm)
#6: Chemical ChemComp-WV8 / (2R,4S)-4-[(2S)-2-phenylbutyl]-1,3-oxazolidin-2-amine


Mass: 218.295 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C13H18N2O / Feature type: SUBJECT OF INVESTIGATION / Comment: agonist*YM

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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

ComponentName: hTA1-Gs-Nb35 complex (Ro5256390) / Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT
Molecular weightUnits: MEGADALTONS / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm) / Strain: Sf9
Buffer solutionpH: 7.4
SpecimenConc.: 1.2 mg/ml / Embedding 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: LEICA EM GP / Cryogen 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: 500 nm
Image recordingElectron dose: 53.88 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.35 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 626370 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00516759
ELECTRON MICROSCOPYf_angle_d0.61830113
ELECTRON MICROSCOPYf_dihedral_angle_d17.0826811
ELECTRON MICROSCOPYf_chiral_restr0.0431304
ELECTRON MICROSCOPYf_plane_restr0.0032517

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