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- PDB-12ag: Spermine-blocked open-state cryo-EM structure of human TRPV6 chan... -

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Basic information

Entry
Database: PDB / ID: 12ag
TitleSpermine-blocked open-state cryo-EM structure of human TRPV6 channel in cNW30 nanodiscs
ComponentsTransient receptor potential cation channel subfamily V member 6
KeywordsMEMBRANE PROTEIN / polyamine / spermine / cancer / cryo-EM / TRP channels / TRPV6 / oncochannel / channel block
Function / homology
Function and homology information


parathyroid hormone secretion / regulation of calcium ion-dependent exocytosis / TRP channels / calcium ion import across plasma membrane / calcium ion homeostasis / calcium channel complex / response to calcium ion / calcium ion transmembrane transport / calcium channel activity / calcium ion transport ...parathyroid hormone secretion / regulation of calcium ion-dependent exocytosis / TRP channels / calcium ion import across plasma membrane / calcium ion homeostasis / calcium channel complex / response to calcium ion / calcium ion transmembrane transport / calcium channel activity / calcium ion transport / calmodulin binding / metal ion binding / identical protein binding / plasma membrane
Similarity search - Function
Transient receptor potential cation channel subfamily V member 6 / Transient receptor potential cation channel subfamily V member 5/6 / Transient receptor potential cation channel subfamily V / Ankyrin repeat / Ankyrin repeat profile. / Ankyrin repeats (3 copies) / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily ...Transient receptor potential cation channel subfamily V member 6 / Transient receptor potential cation channel subfamily V member 5/6 / Transient receptor potential cation channel subfamily V / Ankyrin repeat / Ankyrin repeat profile. / Ankyrin repeats (3 copies) / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
1,2-DIOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / Chem-POV / SPERMINE / CHOLESTEROL HEMISUCCINATE / Transient receptor potential cation channel subfamily V member 6
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.48 Å
AuthorsNeuberger, A. / Sobolevsky, A.I.
Funding support Germany, France, United States, 7items
OrganizationGrant numberCountry
German Research Foundation (DFG)464295817 Germany
Human Frontier Science Program (HFSP) France
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS)AR078814 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS083660 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS107253 United States
National Institutes of Health/National Cancer Institute (NIH/NCI)CA206573 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL181985 United States
CitationJournal: Nat Commun / Year: 2026
Title: Open-channel block of human TRPV6 by polyamine spermine.
Authors: Arthur Neuberger / Irina I Veretenenko / Alexey Shalygin / Yury A Trofimov / Thomas Gudermann / Vladimir Chubanov / Roman G Efremov / Alexander I Sobolevsky /
Abstract: Polyamines are organic cations that are present at sub-millimolar concentrations in the cytoplasm and extracellular fluids and serve as versatile modulators of TRP channels, fine-tuning their ...Polyamines are organic cations that are present at sub-millimolar concentrations in the cytoplasm and extracellular fluids and serve as versatile modulators of TRP channels, fine-tuning their functions in physiological and pathological contexts, including pain, inflammation and cancer. Despite extensive functional studies, the structural basis by which polyamines regulate TRP channels remains unclear. Here, we combine calcium imaging, electrophysiology, cryo-electron microscopy, mutagenesis and molecular dynamics simulations to study regulation of human TRPV6 by polyamine spermine. Our functional experiments demonstrate voltage-dependent block of TRPV6-mediated currents by spermine. Cryo-electron microscopy reveals that spermine binds in the open pore of TRPV6, extending along the pore axis through the selectivity filter and central cavity. Mutagenesis and molecular dynamics simulations confirm the main binding site of spermine in the selectivity filter and suggest a stepwise molecular mechanism of channel block that includes two more binding sites in the pore transiently occupied by spermine. Our findings enrich the knowledge about TRPV6 regulation by endogenous factors and provide details of the ion channel blocking mechanism that can be explored for inhibition of this channel in disease conditions.
History
DepositionMar 23, 2026Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jun 10, 2026Provider: repository / Type: Initial release
Revision 1.0Jun 10, 2026Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Jun 10, 2026Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Jun 10, 2026Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Jun 10, 2026Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Jun 10, 2026Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Transient receptor potential cation channel subfamily V member 6
B: Transient receptor potential cation channel subfamily V member 6
C: Transient receptor potential cation channel subfamily V member 6
D: Transient receptor potential cation channel subfamily V member 6
hetero molecules


Theoretical massNumber of molelcules
Total (without water)298,39133
Polymers280,0784
Non-polymers18,31229
Water724
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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Protein , 1 types, 4 molecules ABCD

#1: Protein
Transient receptor potential cation channel subfamily V member 6 / TrpV6 / CaT-like / CaT-L / Calcium transport protein 1 / CaT1 / Epithelial calcium channel 2 / ECaC2


Mass: 70019.570 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: TRPV6, ECAC2 / Production host: Homo sapiens (human) / References: UniProt: Q9H1D0

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Non-polymers , 5 types, 33 molecules

#2: Chemical
ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C31H50O4
#3: Chemical
ChemComp-PCW / 1,2-DIOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / (Z,Z)-4-HYDROXY-N,N,N-TRIMETHYL-10-OXO-7-[(1-OXO-9-OCTADECENYL)OXY]-3,5,9-TRIOXA-4-PHOSPHAHEPTACOS-18-EN-1-AMINIUM-4-OXIDE


Mass: 787.121 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C44H85NO8P / Comment: DOPC, phospholipid*YM
#4: Chemical
ChemComp-POV / (2S)-3-(hexadecanoyloxy)-2-[(9Z)-octadec-9-enoyloxy]propyl 2-(trimethylammonio)ethyl phosphate / POPC


Mass: 760.076 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C42H82NO8P / Comment: phospholipid*YM
#5: Chemical ChemComp-SPM / SPERMINE


Mass: 202.340 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C10H26N4 / Feature type: SUBJECT OF INVESTIGATION
#6: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY
Has protein modificationN

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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: spermine-blocked open-state homotetrameric human TRPV6
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.08322 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
1150 mMsodium chlorideNaCl1
220 mMtris(hydroxymethyl)aminomethane1
31 mMbeta-Mercaptoethanol1
410 mMspermine1
SpecimenConc.: 2.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: human TRPV6 reconstituted in cNW30 nanodiscs in the presence of spermine
Specimen supportGrid material: GOLD / Grid mesh size: 300 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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm
Image recordingAverage exposure time: 2 sec. / Electron dose: 60 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4487
Image scansWidth: 5760 / Height: 4092

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Processing

EM software
IDNameVersionCategory
1cryoSPARC3.3particle selection
2SerialEMimage acquisition
4cryoSPARC3.3CTF correction
7Cootmodel fitting
9cryoSPARC3.3initial Euler assignment
11cryoSPARC3.3classification
12cryoSPARC3.33D reconstruction
13PHENIXmodel refinement
CTF correctionType: NONE
Particle selectionNum. of particles selected: 2191044
3D reconstructionResolution: 3.48 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 14018 / Symmetry type: POINT
Atomic model buildingSpace: REAL
Atomic model buildingPDB-ID: 7S89

7s89


Accession code: 7S89 / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00821161
ELECTRON MICROSCOPYf_angle_d1.29728628
ELECTRON MICROSCOPYf_dihedral_angle_d16.60712611
ELECTRON MICROSCOPYf_chiral_restr0.0783208
ELECTRON MICROSCOPYf_plane_restr0.0083500

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