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- EMDB-15981: Human apo TRPM8 in a closed state (composite map) -

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Basic information

Entry
Database: EMDB / ID: EMD-15981
TitleHuman apo TRPM8 in a closed state (composite map)
Map datacomposite map created by combining consensus map with four copies of focused pre-MHR MHR1/2 map
Sample
  • Complex: Human TRPM8 in apo form
    • Protein or peptide: Transient receptor potential cation channel subfamily M member 8
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate
  • Ligand: UNDECANE
  • Ligand: SODIUM IONSodium
Keywordscalcium ion channel / cold temperature sensor / MEMBRANE PROTEIN
Function / homology
Function and homology information


ligand-gated calcium channel activity / thermoception / TRP channels / response to cold / calcium ion transmembrane transport / calcium channel activity / intracellular calcium ion homeostasis / positive regulation of cold-induced thermogenesis / membrane raft / external side of plasma membrane ...ligand-gated calcium channel activity / thermoception / TRP channels / response to cold / calcium ion transmembrane transport / calcium channel activity / intracellular calcium ion homeostasis / positive regulation of cold-induced thermogenesis / membrane raft / external side of plasma membrane / endoplasmic reticulum membrane / identical protein binding / plasma membrane
Similarity search - Function
TRPM, SLOG domain / SLOG in TRPM / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Transient receptor potential cation channel subfamily M member 8
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.65 Å
AuthorsPalchevskyi S / Czarnocki-Cieciura M / Vistoli G / Gervasoni S / Nowak E / Beccari AR / Nowotny M / Talarico C
Funding support Poland, 1 items
OrganizationGrant numberCountry
Other government Poland
CitationJournal: Commun Biol / Year: 2023
Title: Structure of human TRPM8 channel.
Authors: Sergii Palchevskyi / Mariusz Czarnocki-Cieciura / Giulio Vistoli / Silvia Gervasoni / Elżbieta Nowak / Andrea R Beccari / Marcin Nowotny / Carmine Talarico /
Abstract: TRPM8 is a non-selective cation channel permeable to both monovalent and divalent cations that is activated by multiple factors, such as temperature, voltage, pressure, and changes in osmolality. It ...TRPM8 is a non-selective cation channel permeable to both monovalent and divalent cations that is activated by multiple factors, such as temperature, voltage, pressure, and changes in osmolality. It is a therapeutic target for anticancer drug development, and its modulators can be utilized for several pathological conditions. Here, we present a cryo-electron microscopy structure of a human TRPM8 channel in the closed state that was solved at 2.7 Å resolution. Our structure comprises the most complete model of the N-terminal pre-melastatin homology region. We also visualized several lipids that are bound by the protein and modeled how the human channel interacts with icilin. Analyses of pore helices in available TRPM structures showed that all these structures can be grouped into different closed, desensitized and open state conformations based on the register of the pore helix S6 which positions particular amino acid residues at the channel constriction.
History
DepositionOct 19, 2022-
Header (metadata) releaseNov 1, 2023-
Map releaseNov 1, 2023-
UpdateNov 1, 2023-
Current statusNov 1, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_15981.map.gz / Format: CCP4 / Size: 282.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationcomposite map created by combining consensus map with four copies of focused pre-MHR MHR1/2 map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.82 Å/pix.
x 420 pix.
= 344.4 Å
0.82 Å/pix.
x 420 pix.
= 344.4 Å
0.82 Å/pix.
x 420 pix.
= 344.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.82 Å
Density
Contour LevelBy AUTHOR: 0.6
Minimum - Maximum0.0 - 5.208554
Average (Standard dev.)0.007377137 (±0.07505462)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions420420420
Spacing420420420
CellA=B=C: 344.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: raw consensus map

Fileemd_15981_additional_1.map
Annotationraw consensus map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : Human TRPM8 in apo form

EntireName: Human TRPM8 in apo form
Components
  • Complex: Human TRPM8 in apo form
    • Protein or peptide: Transient receptor potential cation channel subfamily M member 8
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate
  • Ligand: UNDECANE
  • Ligand: SODIUM IONSodium

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Supramolecule #1: Human TRPM8 in apo form

SupramoleculeName: Human TRPM8 in apo form / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 510 KDa

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

MacromoleculeName: Transient receptor potential cation channel subfamily M member 8
type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 127.904719 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: SNSFRAARLS MRNRRNDTLD STRTLYSSAS RSTDLSYSES DLVNFIQANF KKRECVFFTK DSKATENVCK CGYAQSQHME GTQINQSEK WNYKKHTKEF PTDAFGDIQF ETLGKKGKYI RLSCDTDAEI LYELLTQHWH LKTPNLVISV TGGAKNFALK P RMRKIFSR ...String:
SNSFRAARLS MRNRRNDTLD STRTLYSSAS RSTDLSYSES DLVNFIQANF KKRECVFFTK DSKATENVCK CGYAQSQHME GTQINQSEK WNYKKHTKEF PTDAFGDIQF ETLGKKGKYI RLSCDTDAEI LYELLTQHWH LKTPNLVISV TGGAKNFALK P RMRKIFSR LIYIAQSKGA WILTGGTHYG LMKYIGEVVR DNTISRSSEE NIVAIGIAAW GMVSNRDTLI RNCDAEGYFL AQ YLMDDFT RDPLYILDNN HTHLLLVDNG CHGHPTVEAK LRNQLEKYIS ERTIQDSNYG GKIPIVCFAQ GGGKETLKAI NTS IKNKIP CVVVEGSGQI ADVIASLVEV EDALTSSAVK EKLVRFLPRT VSRLPEEETE SWIKWLKEIL ECSHLLTVIK MEEA GDEIV SNAISYALYK AFSTSEQDKD NWNGQLKLLL EWNQLDLAND EIFTNDRRWE SADLQEVMFT ALIKDRPKFV RLFLE NGLN LRKFLTHDVL TELFSNHFST LVYRNLQIAK NSYNDALLTF VWKLVANFRR GFRKEDRNGR DEMDIELHDV SPITRH PLQ ALFIWAILQN KKELSKVIWE QTRGCTLAAL GASKLLKTLA KVKNDINAAG ESEELANEYE TRAVELFTEC YSSDEDL AE QLLVYSCEAW GGSNCLELAV EATDQHFIAQ PGVQNFLSKQ WYGEISRDTK NWKIILCLFI IPLVGCGFVS FRKKPVDK H KKLLWYYVAF FTSPFVVFSW NVVFYIAFLL LFAYVLLMDF HSVPHPPELV LYSLVFVLFC DEVRQWYVNG VNYFTDLWN VMDTLGLFYF IAGIVFRLHS SNKSSLYSGR VIFCLDYIIF TLRLIHIFTV SRNLGPKIIM LQRMLIDVFF FLFLFAVWMV AFGVARQGI LRQNEQRWRW IFRSVIYEPY LAMFGQVPSD VDGTTYDFAH CTFTGNESKP LCVELDEHNL PRFPEWITIP L VCIYMLST NILLVNLLVA MFGYTVGTVQ ENNDQVWKFQ RYFLVQEYCS RLNIPFPFIV FAYFYMVVKK CFKCCCKEKN ME SSVCCFK NEDNETLAWE GVMKENYLVK INTKANDTSE EMRHRFRQLD TKLNDLKGLL KEIANKIK

UniProtKB: Transient receptor potential cation channel subfamily M member 8

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Macromolecule #2: CHOLESTEROL HEMISUCCINATE

MacromoleculeName: CHOLESTEROL HEMISUCCINATE / type: ligand / ID: 2 / Number of copies: 8 / Formula: Y01
Molecular weightTheoretical: 486.726 Da
Chemical component information

ChemComp-Y01:
CHOLESTEROL HEMISUCCINATE

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Macromolecule #3: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(tri...

MacromoleculeName: (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate
type: ligand / ID: 3 / Number of copies: 4 / Formula: POV
Molecular weightTheoretical: 760.076 Da
Chemical component information

ChemComp-POV:
(2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate / phospholipid*YM / POPC

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Macromolecule #4: UNDECANE

MacromoleculeName: UNDECANE / type: ligand / ID: 4 / Number of copies: 4 / Formula: UND
Molecular weightTheoretical: 156.308 Da
Chemical component information

ChemComp-UND:
UNDECANE / Undecane

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Macromolecule #5: SODIUM ION

MacromoleculeName: SODIUM ION / type: ligand / ID: 5 / Number of copies: 8
Molecular weightTheoretical: 22.99 Da

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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

Concentration0.5 mg/mL
BufferpH: 7.4
GridModel: UltrAuFoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE
Details: The grid was glow-discharged from both sides prior to vitrification
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
Detailsprotein solubilized by LMNG

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.7 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 105000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 7918 / Average electron dose: 41.42 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1239855
Startup modelType of model: OTHER / Details: cryoSPARC Ab-Initio reconstruction
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final 3D classificationNumber classes: 6 / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.65 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2)
Details: composite map created by combining consensus map with four copies of focused pre-MHR + MHR1/2 map
Number images used: 110176

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8bdc:
Human apo TRPM8 in a closed state (composite map)

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