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- PDB-8e4q: The closed C0-state flycatcher TRPM8 structure in complex with PI... -

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Basic information

Entry
Database: PDB / ID: 8e4q
TitleThe closed C0-state flycatcher TRPM8 structure in complex with PI(4,5)P2
ComponentsTransient receptor potential cation channel subfamily M member 8
KeywordsMEMBRANE PROTEIN / TRPM8 / menthol receptor / cold receptor / cold sensor / PI(4 / 5)P2 / cooling agonists / temperature sensing / ion channel / sensory transduction / transient receptor potential ion channel
Function / homology
Function and homology information


monoatomic ion channel activity / identical protein binding / membrane
Similarity search - Function
TRPM, SLOG domain / : / SLOG in TRPM / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Chem-PIO / Transient receptor potential cation channel subfamily M member 8
Similarity search - Component
Biological speciesFicedula albicollis (Collared flycatcher)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.51 Å
AuthorsYin, Y. / Zhang, F. / Feng, S. / Butay, K.J. / Borgnia, M.J. / Im, W. / Lee, S.-Y.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Eye Institute (NIH/NEI)R01EY031698 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R35NS097241 United States
CitationJournal: Science / Year: 2022
Title: Activation mechanism of the mouse cold-sensing TRPM8 channel by cooling agonist and PIP.
Authors: Ying Yin / Feng Zhang / Shasha Feng / Kevin John Butay / Mario J Borgnia / Wonpil Im / Seok-Yong Lee /
Abstract: The transient receptor potential melastatin 8 (TRPM8) channel is the primary molecular transducer responsible for the cool sensation elicited by menthol and cold in mammals. TRPM8 activation is ...The transient receptor potential melastatin 8 (TRPM8) channel is the primary molecular transducer responsible for the cool sensation elicited by menthol and cold in mammals. TRPM8 activation is controlled by cooling compounds together with the membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP). Our knowledge of cold sensation and the therapeutic potential of TRPM8 for neuroinflammatory diseases and pain will be enhanced by understanding the structural basis of cooling agonist- and PIP-dependent TRPM8 activation. We present cryo-electron microscopy structures of mouse TRPM8 in closed, intermediate, and open states along the ligand- and PIP-dependent gating pathway. Our results uncover two discrete agonist sites, state-dependent rearrangements in the gate positions, and a disordered-to-ordered transition of the gate-forming S6-elucidating the molecular basis of chemically induced cool sensation in mammals.
History
DepositionAug 18, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 26, 2022Provider: repository / Type: Initial release
Revision 1.1Jun 12, 2024Group: Data collection / Refinement description
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / em_3d_fitting_list / pdbx_initial_refinement_model
Item: _em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id ..._em_3d_fitting_list.accession_code / _em_3d_fitting_list.initial_refinement_model_id / _em_3d_fitting_list.source_name / _em_3d_fitting_list.type

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Transient receptor potential cation channel subfamily M member 8
B: Transient receptor potential cation channel subfamily M member 8
D: Transient receptor potential cation channel subfamily M member 8
C: Transient receptor potential cation channel subfamily M member 8
hetero molecules


Theoretical massNumber of molelcules
Total (without water)528,3608
Polymers525,3744
Non-polymers2,9864
Water00
1


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

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Components

#1: Protein
Transient receptor potential cation channel subfamily M member 8


Mass: 131343.406 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Ficedula albicollis (Collared flycatcher)
Production host: Homo sapiens (human) / References: UniProt: U3JD03
#2: Chemical
ChemComp-PIO / [(2R)-2-octanoyloxy-3-[oxidanyl-[(1R,2R,3S,4R,5R,6S)-2,3,6-tris(oxidanyl)-4,5-diphosphonooxy-cyclohexyl]oxy-phosphoryl]oxy-propyl] octanoate / dioctanoyl l-alpha-phosphatidyl-d-myo-inositol 4,5-diphosphate


Mass: 746.566 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C25H49O19P3
Has ligand of interestY
Sequence detailsAs per the authors, the old version of Uniprot U3JD03 was used for gene synthesis of the sample sequence

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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: Transient receptor potential cation channel subfamily M member 8
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Ficedula albicollis (Collared flycatcher)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
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: 277.15 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 FIELD / Nominal magnification: 75000 X / Nominal defocus max: 3000 nm / Nominal defocus min: 1250 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 42 e/Å2 / Detector mode: COUNTING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4283

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
EM software
IDNameCategory
2EPUimage acquisition
4GctfCTF correction
5RELIONCTF correction
8Cootmodel fitting
10RELIONinitial Euler assignment
11cryoSPARCfinal Euler assignment
13cryoSPARC3D reconstruction
14PHENIXmodel refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1308147
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 3.51 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 51879 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Atomic model buildingPDB-ID: 6NR2

6nr2


Accession code: 6NR2 / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00428176
ELECTRON MICROSCOPYf_angle_d0.56238368
ELECTRON MICROSCOPYf_dihedral_angle_d13.0449352
ELECTRON MICROSCOPYf_chiral_restr0.0384484
ELECTRON MICROSCOPYf_plane_restr0.0044708

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