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- PDB-8pm2: Structure of the murine trace amine-associated receptor TAAR7f bo... -

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Basic information

Entry
Database: PDB / ID: 8pm2
TitleStructure of the murine trace amine-associated receptor TAAR7f bound to N,N-dimethylcyclohexylamine (DMCH) in complex with mini-Gs trimeric G protein
Components
  • (Guanine nucleotide-binding protein ...) x 3
  • Nanobody 35
  • Trace amine-associated receptor 7f
KeywordsMEMBRANE PROTEIN / trace-amine associated receptor / TAAR / mTAAR7f / GPCR / receptor / G protein
Function / homology
Function and homology information


trace-amine receptor activity / sensory perception of chemical stimulus / 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 / G protein-coupled acetylcholine receptor signaling pathway / G-protein activation / Activation of G protein gated Potassium channels ...trace-amine receptor activity / sensory perception of chemical stimulus / 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 / G protein-coupled acetylcholine receptor signaling pathway / 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 / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / G beta:gamma signalling through BTK / ADP signalling through P2Y purinoceptor 12 / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / photoreceptor disc membrane / Glucagon-type ligand receptors / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / cellular response to catecholamine stimulus / ADORA2B mediated anti-inflammatory cytokines production / sensory perception of taste / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / adenylate cyclase-activating dopamine receptor signaling pathway / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / GPER1 signaling / cellular response to prostaglandin E stimulus / Inactivation, recovery and regulation of the phototransduction cascade / G-protein beta-subunit binding / heterotrimeric G-protein complex / G alpha (12/13) signalling events / extracellular vesicle / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / GTPase binding / retina development in camera-type eye / Ca2+ pathway / phospholipase C-activating G protein-coupled receptor signaling pathway / 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 / G protein-coupled receptor signaling pathway / lysosomal membrane / GTPase activity / synapse / protein-containing complex binding / signal transduction / extracellular exosome / membrane / plasma membrane / cytosol / cytoplasm
Similarity search - Function
Trace amine associated receptor family / : / Serpentine type 7TM GPCR chemoreceptor Srsx / 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 ...Trace amine associated receptor family / : / Serpentine type 7TM GPCR chemoreceptor Srsx / 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. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
~{N},~{N}-dimethylcyclohexanamine / CHOLESTEROL HEMISUCCINATE / 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 / Trace amine-associated receptor 7f
Similarity search - Component
Biological speciesHomo sapiens (human)
Lama glama (llama)
Mus musculus (house mouse)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.92 Å
AuthorsGusach, A. / Lee, Y. / Edwards, P.C. / Huang, F. / Weyand, S.N. / Tate, C.G.
Funding support United Kingdom, 1items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_U105197215 United Kingdom
CitationJournal: bioRxiv / Year: 2023
Title: Molecular recognition of an aversive odorant by the murine trace amine-associated receptor TAAR7f.
Authors: Anastasiia Gusach / Yang Lee / Armin Nikpour Khoshgrudi / Elizaveta Mukhaleva / Ning Ma / Eline J Koers / Qingchao Chen / Patricia C Edwards / Fanglu Huang / Jonathan Kim / Filippo Mancia / ...Authors: Anastasiia Gusach / Yang Lee / Armin Nikpour Khoshgrudi / Elizaveta Mukhaleva / Ning Ma / Eline J Koers / Qingchao Chen / Patricia C Edwards / Fanglu Huang / Jonathan Kim / Filippo Mancia / Dmitry B Verprintsev / Nagarajan Vaidehi / Simone N Weyand / Christopher G Tate /
Abstract: There are two main families of G protein-coupled receptors that detect odours in humans, the odorant receptors (ORs) and the trace amine-associated receptors (TAARs). Their amino acid sequences are ...There are two main families of G protein-coupled receptors that detect odours in humans, the odorant receptors (ORs) and the trace amine-associated receptors (TAARs). Their amino acid sequences are distinct, with the TAARs being most similar to the aminergic receptors such as those activated by adrenaline, serotonin and histamine. To elucidate the structural determinants of ligand recognition by TAARs, we have determined the cryo-EM structure of a murine receptor, mTAAR7f, coupled to the heterotrimeric G protein G and bound to the odorant N,N-dimethylcyclohexylamine (DMCH) to an overall resolution of 2.9 Å. DMCH is bound in a hydrophobic orthosteric binding site primarily through van der Waals interactions and a strong charge-charge interaction between the tertiary amine of the ligand and an aspartic acid residue. This site is distinct and non-overlapping with the binding site for the odorant propionate in the odorant receptor OR51E2. The structure, in combination with mutagenesis data and molecular dynamics simulations suggests that the activation of the receptor follows a similar pathway to that of the β-adrenoceptors, with the significant difference that DMCH interacts directly with one of the main activation microswitch residues.
History
DepositionJun 28, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Aug 9, 2023Provider: repository / Type: Initial release
Revision 2.0Jul 17, 2024Group: Advisory / Atomic model ...Advisory / Atomic model / Author supporting evidence / Data collection / Derived calculations / Refinement description / Structure summary
Category: atom_site / chem_comp_atom ...atom_site / chem_comp_atom / chem_comp_bond / em_admin / em_imaging / entity / pdbx_entity_instance_feature / pdbx_entity_nonpoly / pdbx_nonpoly_scheme / pdbx_poly_seq_scheme / pdbx_struct_sheet_hbond / pdbx_unobs_or_zero_occ_atoms / pdbx_unobs_or_zero_occ_residues / pdbx_validate_close_contact / pdbx_validate_torsion / refine / refine_hist / refine_ls_restr / refine_ls_shell / struct_conf / struct_conn / struct_sheet / struct_sheet_order / struct_sheet_range
Item: _em_admin.last_update / _em_imaging.nominal_defocus_max ..._em_admin.last_update / _em_imaging.nominal_defocus_max / _entity.formula_weight / _entity.pdbx_description / _pdbx_entity_nonpoly.comp_id / _pdbx_entity_nonpoly.name / _pdbx_nonpoly_scheme.auth_mon_id / _pdbx_nonpoly_scheme.auth_seq_num / _pdbx_nonpoly_scheme.mon_id / _pdbx_nonpoly_scheme.pdb_mon_id / _pdbx_poly_seq_scheme.auth_mon_id / _pdbx_poly_seq_scheme.auth_seq_num / _pdbx_poly_seq_scheme.pdb_mon_id / _refine.B_iso_mean / _refine.correlation_coeff_Fo_to_Fc / _refine.details / _refine.ls_R_factor_R_work / _refine.ls_R_factor_obs / _refine.ls_d_res_high / _refine.ls_d_res_low / _refine.ls_number_reflns_obs / _refine.ls_percent_reflns_obs / _refine.overall_SU_B / _refine.overall_SU_ML / _refine.pdbx_overall_ESU_R / _refine.pdbx_stereochemistry_target_values / _refine.solvent_model_details / _struct_conn.pdbx_dist_value / _struct_sheet.number_strands
Revision 2.1Aug 7, 2024Group: Data collection / Category: em_admin / Item: _em_admin.last_update
Revision 2.2Oct 23, 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: 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 35
R: Trace amine-associated receptor 7f
hetero molecules


Theoretical massNumber of molelcules
Total (without water)132,7057
Polymers132,0915
Non-polymers6142
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area12000 Å2
ΔGint-68 kcal/mol
Surface area41780 Å2

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Components

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

#1: Protein Guanine nucleotide-binding protein G(s) subunit alpha isoforms short


Mass: 28964.734 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 (bacteria) / Variant (production host): CodonPlus(DE3)-RIL
#2: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1


Mass: 37342.785 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P62873
#3: 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 / Mutation: C68S
Source method: isolated from a genetically manipulated source
Details: There is an engineered mutation C68S introduced / Source: (gene. exp.) Homo sapiens (human) / Gene: GNG2 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P59768

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Antibody / Protein , 2 types, 2 molecules NR

#4: Antibody Nanobody 35


Mass: 16926.076 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 (bacteria) / Variant (production host): CodonPlus(DE3)-RIL
#5: Protein Trace amine-associated receptor 7f / TaR-7f / Trace amine receptor 7f / mTaar7f


Mass: 41012.539 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: mTAAR7f sequence contains cleaved protease sites: TEV (S residue on the N terminus) and HRV-3C (C terminus)
Source: (gene. exp.) Mus musculus (house mouse) / Gene: Taar7f / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q5QD08

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Non-polymers , 2 types, 2 molecules

#6: Chemical ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C31H50O4
#7: Chemical ChemComp-8IA / ~{N},~{N}-dimethylcyclohexanamine


Mass: 127.227 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C8H17N

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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: A complex of mouse trace-amine associated receptor 7f solubilized in LMNG/CHS bound to N,N-dimethylcyclohexylamine and coupled to: Engineered guanine nucleotide-binding protein G(s) subunit ...Name: A complex of mouse trace-amine associated receptor 7f solubilized in LMNG/CHS bound to N,N-dimethylcyclohexylamine and coupled to: Engineered guanine nucleotide-binding protein G(s) subunit alpha isoform short + Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 + Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 + Nanobody 35
Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPES/KOHC8H17KN2O4S1
2150 mMPotassium chlorideKCL1
310 mMSodium chlorideNaCl1
410 mMMagnesium chlorideMgCl21
50.01 %Lauryl Maltose Neopentyl Glycol1
60.002 %Cholesteryl Hemisuccinate1
76.7 mMN,N-dimethylcyclohexylamine1
SpecimenConc.: 0.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: Forward Power of 38 W, Reflected Power of 2W; Fischione
Grid material: GOLD / Grid mesh size: 400 divisions/in. / Grid type: UltrAuFoil 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: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELD / Nominal magnification: 96000 X / Calibrated magnification: 96000 X / Nominal defocus max: 2400 nm / Nominal defocus min: 800 nm / Calibrated defocus min: 2400 nm / Calibrated defocus max: 2400 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 7.84 sec. / Electron dose: 55 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 11157
Image scansWidth: 4096 / Height: 4096

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4.1.1+patch23011029particle selection
4cryoSPARC4.1.1+patch23011029CTF correction
7cryoSPARC4.1.1+patch23011029model fitting
9cryoSPARC4.1.1+patch23011029initial Euler assignment
10cryoSPARC4.1.1+patch23011029final Euler assignment
11cryoSPARC4.1.1+patch23011029classification
12cryoSPARC4.1.1+patch230110293D reconstruction
13cryoSPARC4.1.1+patch23011029model refinement
CTF correctionDetails: Done with cryoSPARC patch motion correction / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 9199357 / Details: Particles were heavily over-picked
3D reconstructionResolution: 2.92 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 172639 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL
Atomic model buildingDetails: Initial models of heterotrimeric mini-Gs and Nb35 were sourced from PDB 7T9I. A de novo model of TAAR7f was generated from the focused map and protein sequence using ModelAngelo
Source name: Other / Type: other
RefinementResolution: 2.92→2.92 Å / Cor.coef. Fo:Fc: 0.836 / SU B: 13.865 / SU ML: 0.249 / ESU R: 0.346
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN USED IF PRESENT IN THE INPUT
RfactorNum. reflection% reflection
Rwork0.39743 --
obs0.39743 109353 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 104.792 Å2
Refinement stepCycle: 1 / Total: 7624
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0070.0117797
ELECTRON MICROSCOPYr_bond_other_d00.0167194
ELECTRON MICROSCOPYr_angle_refined_deg1.2141.64310621
ELECTRON MICROSCOPYr_angle_other_deg0.421.57216477
ELECTRON MICROSCOPYr_dihedral_angle_1_deg5.1435998
ELECTRON MICROSCOPYr_dihedral_angle_2_deg0.4160.15547
ELECTRON MICROSCOPYr_dihedral_angle_3_deg14.027101152
ELECTRON MICROSCOPYr_dihedral_angle_4_deg
ELECTRON MICROSCOPYr_chiral_restr0.0550.21225
ELECTRON MICROSCOPYr_gen_planes_refined0.0060.029335
ELECTRON MICROSCOPYr_gen_planes_other0.0010.021907
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it11.169.9324016
ELECTRON MICROSCOPYr_mcbond_other11.1599.9324016
ELECTRON MICROSCOPYr_mcangle_it17.38817.8355006
ELECTRON MICROSCOPYr_mcangle_other17.38717.8375007
ELECTRON MICROSCOPYr_scbond_it12.72510.9753781
ELECTRON MICROSCOPYr_scbond_other12.72410.9753782
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other20.46719.7215616
ELECTRON MICROSCOPYr_long_range_B_refined30.848118.3331515
ELECTRON MICROSCOPYr_long_range_B_other30.848118.3331516
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 2.8→2.873 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork1.284 8084 -
obs--100 %

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