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- EMDB-41219: Wildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2... -

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Basic information

Entry
Database: EMDB / ID: EMD-41219
TitleWildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2 and econazole
Map dataTRPV5 PIP2 ECN
Sample
  • Complex: Tetramer of wildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2 and econazole
    • Protein or peptide: Transient receptor potential cation channel subfamily V member 5
  • Ligand: ERGOSTEROL
  • Ligand: 1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE
  • Ligand: 1-[(2S)-2-[(4-CHLOROBENZYL)OXY]-2-(2,4-DICHLOROPHENYL)ETHYL]-1H-IMIDAZOLE
KeywordsTRPV5 / TRP channel / PI(4 / 5)P2 / econazole / MEMBRANE PROTEIN
Function / homology
Function and homology information


regulation of urine volume / calcium ion import across plasma membrane / calcium ion homeostasis / calcium ion transmembrane transport / calcium channel activity / calcium ion transport / protein homotetramerization / calmodulin binding / apical plasma membrane / identical protein binding ...regulation of urine volume / calcium ion import across plasma membrane / calcium ion homeostasis / calcium ion transmembrane transport / calcium channel activity / calcium ion transport / protein homotetramerization / calmodulin binding / apical plasma membrane / identical protein binding / metal ion binding / plasma membrane
Similarity search - Function
Transient receptor potential cation channel subfamily V member 5 / Transient receptor potential cation channel subfamily V member 5/6 / Transient receptor potential cation channel subfamily V / Ankyrin repeat / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily ...Transient receptor potential cation channel subfamily V member 5 / Transient receptor potential cation channel subfamily V member 5/6 / Transient receptor potential cation channel subfamily V / Ankyrin repeat / Ankyrin repeats (3 copies) / Ankyrin repeat profile. / Ankyrin repeat region circular profile. / ankyrin repeats / Ankyrin repeat / Ankyrin repeat-containing domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Transient receptor potential cation channel subfamily V member 5
Similarity search - Component
Biological speciesOryctolagus cuniculus (rabbit)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.86 Å
AuthorsDe Jesus-Perez JJ / Fluck EC / Moiseenkova-Bell VY
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM144120 United States
CitationJournal: Structure / Year: 2024
Title: Structural mechanism of TRPV5 inhibition by econazole.
Authors: José J De Jesús-Pérez / Matthew Gabrielle / Sumiyya Raheem / Edwin C Fluck / Tibor Rohacs / Vera Y Moiseenkova-Bell /
Abstract: The calcium-selective TRPV5 channel activated by phosphatidylinositol 4,5-bisphosphate [PI(4,5)P] is involved in calcium homeostasis. Recently, cryoelectron microscopy (cryo-EM) provided molecular ...The calcium-selective TRPV5 channel activated by phosphatidylinositol 4,5-bisphosphate [PI(4,5)P] is involved in calcium homeostasis. Recently, cryoelectron microscopy (cryo-EM) provided molecular details of TRPV5 modulation by exogenous and endogenous molecules. However, the details of TRPV5 inhibition by the antifungal agent econazole (ECN) remain elusive due to the low resolution of the currently available structure. In this study, we employ cryo-EM to comprehensively examine how the ECN inhibits TRPV5. By combining our structural findings with site-directed mutagenesis, calcium measurements, electrophysiology, and molecular dynamics simulations, we determined that residues F472 and L475 on the S4 helix, along with residue W495 on the S5 helix, collectively constitute the ECN-binding site. Additionally, the structure of TRPV5 in the presence of ECN and PI(4,5)P, which does not show the bound activator, reveals a potential inhibition mechanism in which ECN competes with PI(4,5)P, preventing the latter from binding, and ultimately pore closure.
History
DepositionJul 7, 2023-
Header (metadata) releaseJan 10, 2024-
Map releaseJan 10, 2024-
UpdateFeb 14, 2024-
Current statusFeb 14, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41219.png

Downloads & links

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Map

FileDownload / File: emd_41219.map.gz / Format: CCP4 / Size: 91.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTRPV5 PIP2 ECN
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 288 pix.
= 239.04 Å		0.83 Å/pix.
x 288 pix.
= 239.04 Å		0.83 Å/pix.
x 288 pix.
= 239.04 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.16
Minimum - Maximum-0.8246608 - 1.452503
Average (Standard dev.)0.0032156268 (±0.04743066)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions288288288
Spacing288288288
CellA=B=C: 239.04 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: TRPV5 PIP2 ECN half1

Fileemd_41219_half_map_1.map
AnnotationTRPV5 PIP2 ECN half1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: TRPV5 PIP2 ECN half2

Fileemd_41219_half_map_2.map
AnnotationTRPV5 PIP2 ECN half2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Tetramer of wildtype rabbit TRPV5 into nanodiscs in the presence ...

EntireName: Tetramer of wildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2 and econazole
Components
  • Complex: Tetramer of wildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2 and econazole
    • Protein or peptide: Transient receptor potential cation channel subfamily V member 5
  • Ligand: ERGOSTEROL
  • Ligand: 1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE
  • Ligand: 1-[(2S)-2-[(4-CHLOROBENZYL)OXY]-2-(2,4-DICHLOROPHENYL)ETHYL]-1H-IMIDAZOLE

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Supramolecule #1: Tetramer of wildtype rabbit TRPV5 into nanodiscs in the presence ...

SupramoleculeName: Tetramer of wildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2 and econazole
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Molecular weightTheoretical: 335.14 KDa

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Macromolecule #1: Transient receptor potential cation channel subfamily V member 5

MacromoleculeName: Transient receptor potential cation channel subfamily V member 5
type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Oryctolagus cuniculus (rabbit)
Molecular weightTheoretical: 83.784586 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MGACPPKAKG PWAQLQKLLI SWPVGEQDWE QYRDRVNMLQ QERIRDSPLL QAAKENDLRL LKILLLNQSC DFQQRGAVGE TALHVAALY DNLEAATLLM EAAPELAKEP ALCEPFVGQT ALHIAVMNQN LNLVRALLAR GASVSARATG AAFRRSPHNL I YYGEHPLS ...String:
MGACPPKAKG PWAQLQKLLI SWPVGEQDWE QYRDRVNMLQ QERIRDSPLL QAAKENDLRL LKILLLNQSC DFQQRGAVGE TALHVAALY DNLEAATLLM EAAPELAKEP ALCEPFVGQT ALHIAVMNQN LNLVRALLAR GASVSARATG AAFRRSPHNL I YYGEHPLS FAACVGSEEI VRLLIEHGAD IRAQDSLGNT VLHILILQPN KTFACQMYNL LLSYDEHSDH LQSLELVPNH QG LTPFKLA GVEGNTVMFQ HLMQKRKHVQ WTCGPLTSTL YDLTEIDSWG EELSFLELVV SSKKREARQI LEQTPVKELV SFK WKKYGR PYFCVLASLY ILYMICFTTC CIYRPLKLRD DNRTDPRDIT ILQQKLLQEA YVTHQDNIRL VGELVTVTGA VIIL LLEIP DIFRVGASRY FGQTILGGPF HVIIITYASL VLLTMVMRLT NMNGEVVPLS FALVLGWCSV MYFARGFQML GPFTI MIQK MIFGDLMRFC WLMAVVILGF ASAFHITFQT EDPNNLGEFS DYPTALFSTF ELFLTIIDGP ANYSVDLPFM YCITYA AFA IIATLLMLNL FIAMMGDTHW RVAQERDELW RAQVVATTVM LERKMPRFLW PRSGICGYEY GLGDRWFLRV ENHHDQN PL RVLRYVEAFK CSDKEDGQEQ LSEKRPSTVE SGMLSRASVA FQTPSLSRTT SQSSNSHRGW EILRRNTLGH LNLGLDLG E GDGEEVYHFT ETSQVAPA

UniProtKB: Transient receptor potential cation channel subfamily V member 5

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Macromolecule #2: ERGOSTEROL

MacromoleculeName: ERGOSTEROL / type: ligand / ID: 2 / Number of copies: 8 / Formula: ERG
Molecular weightTheoretical: 396.648 Da
Chemical component information

ChemComp-ERG:
ERGOSTEROL

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Macromolecule #3: 1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE

MacromoleculeName: 1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / type: ligand / ID: 3 / Number of copies: 8 / Formula: CPL
Molecular weightTheoretical: 758.06 Da
Chemical component information

ChemComp-CPL:
1-PALMITOYL-2-LINOLEOYL-SN-GLYCERO-3-PHOSPHOCHOLINE / phospholipid*YM

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Macromolecule #4: 1-[(2S)-2-[(4-CHLOROBENZYL)OXY]-2-(2,4-DICHLOROPHENYL)ETHYL]-1H-I...

MacromoleculeName: 1-[(2S)-2-[(4-CHLOROBENZYL)OXY]-2-(2,4-DICHLOROPHENYL)ETHYL]-1H-IMIDAZOLE
type: ligand / ID: 4 / Number of copies: 4 / Formula: ECN
Molecular weightTheoretical: 381.684 Da
Chemical component information

ChemComp-ECN:
1-[(2S)-2-[(4-CHLOROBENZYL)OXY]-2-(2,4-DICHLOROPHENYL)ETHYL]-1H-IMIDAZOLE

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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

BufferpH: 8
VitrificationCryogen name: ETHANE / Chamber humidity: 100 %

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 11346 / Average electron dose: 41.6 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1316241
Startup modelType of model: EMDB MAP
EMDB ID:

25716

Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.86 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.0.1) / Number images used: 101063
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0.1)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: FLEXIBLE FIT
Output model

PDB-8tf4:
Wildtype rabbit TRPV5 into nanodiscs in the presence of PI(4,5)P2 and econazole

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