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- EMDB-7435: Cryo-EM structure of mouse TPC1 channel in the PtdIns(3,5)P2-boun... -

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Basic information

Entry
Database: EMDB / ID: EMD-7435
TitleCryo-EM structure of mouse TPC1 channel in the PtdIns(3,5)P2-bound state
Map dataTPC1 channel in the PtdIns(3,5)P2-bound state
Sample
  • Organelle or cellular component: PtdIns(3,5)P2-bound mouse TPC1
    • Protein or peptide: Two pore calcium channel protein 1
  • Ligand: (2R)-3-{[(S)-hydroxy{[(1S,2R,3R,4S,5S,6R)-2,4,6-trihydroxy-3,5-bis(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dioctanoate
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose
Function / homology
Function and homology information


endolysosome / intracellularly phosphatidylinositol-3,5-bisphosphate-gated monatomic cation channel activity / ligand-gated sodium channel activity / NAADP-sensitive calcium-release channel activity / voltage-gated monoatomic ion channel activity / Stimuli-sensing channels / voltage-gated sodium channel activity / regulation of monoatomic ion transmembrane transport / phosphatidylinositol-3,5-bisphosphate binding / syntaxin binding ...endolysosome / intracellularly phosphatidylinositol-3,5-bisphosphate-gated monatomic cation channel activity / ligand-gated sodium channel activity / NAADP-sensitive calcium-release channel activity / voltage-gated monoatomic ion channel activity / Stimuli-sensing channels / voltage-gated sodium channel activity / regulation of monoatomic ion transmembrane transport / phosphatidylinositol-3,5-bisphosphate binding / syntaxin binding / sodium ion transmembrane transport / positive regulation of autophagy / endocytosis involved in viral entry into host cell / recycling endosome membrane / early endosome membrane / endosome membrane / lysosomal membrane / protein homodimerization activity / membrane / identical protein binding
Similarity search - Function
Two pore channel protein 1 / Voltage-dependent channel domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Two pore calcium channel protein 1
Similarity search - Component
Biological speciesMus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsShe J / Guo J / Chen Q / Bai X / Jiang Y
Funding support United States, 5 items
OrganizationGrant numberCountry
National Institutes of Health/National Center for Research Resources (NIH/NCRR)GM079179 United States
Howard Hughes Medical Institute (HHMI) United States
Welch FoundationI-1578 United States
Virginia Murchison Linthicum Scholar in Medical Research fund United States
Cancer Prevention and Research Institute of Texas (CPRIT) United States
CitationJournal: Nature / Year: 2018
Title: Structural insights into the voltage and phospholipid activation of the mammalian TPC1 channel.
Authors: Ji She / Jiangtao Guo / Qingfeng Chen / Weizhong Zeng / Youxing Jiang / Xiao-Chen Bai /
Abstract: The organellar two-pore channel (TPC) functions as a homodimer, in which each subunit contains two homologous Shaker-like six-transmembrane (6-TM)-domain repeats. TPCs belong to the voltage-gated ion ...The organellar two-pore channel (TPC) functions as a homodimer, in which each subunit contains two homologous Shaker-like six-transmembrane (6-TM)-domain repeats. TPCs belong to the voltage-gated ion channel superfamily and are ubiquitously expressed in animals and plants. Mammalian TPC1 and TPC2 are localized at the endolysosomal membrane, and have critical roles in regulating the physiological functions of these acidic organelles. Here we present electron cryo-microscopy structures of mouse TPC1 (MmTPC1)-a voltage-dependent, phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P)-activated Na-selective channel-in both the apo closed state and ligand-bound open state. Combined with functional analysis, these structures provide comprehensive structural insights into the selectivity and gating mechanisms of mammalian TPC channels. The channel has a coin-slot-shaped ion pathway in the filter that defines the selectivity of mammalian TPCs. Only the voltage-sensing domain from the second 6-TM domain confers voltage dependence on MmTPC1. Endolysosome-specific PtdIns(3,5)P binds to the first 6-TM domain and activates the channel under conditions of depolarizing membrane potential. Structural comparisons between the apo and PtdIns(3,5)P-bound structures show the interplay between voltage and ligand in channel activation. These MmTPC1 structures reveal lipid binding and regulation in a 6-TM voltage-gated channel, which is of interest in light of the emerging recognition of the importance of phosphoinositide regulation of ion channels.
History
DepositionJan 25, 2018-
Header (metadata) releaseApr 4, 2018-
Map releaseApr 4, 2018-
UpdateJul 29, 2020-
Current statusJul 29, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0446
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0446
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6c9a
  • Surface level: 0.0446
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7435.png

Downloads & links

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Map

FileDownload / File: emd_7435.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTPC1 channel in the PtdIns(3,5)P2-bound state
Voxel sizeX=Y=Z: 1.07 Å
Density
Contour LevelBy EMDB: 0.0446 / Movie #1: 0.0446
Minimum - Maximum-0.1109281 - 0.21509735
Average (Standard dev.)-0.00006319238 (±0.0072761234)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 256.80002 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.071.071.07
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z256.800256.800256.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.1110.215-0.000

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Supplemental data

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Sample components

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Entire : PtdIns(3,5)P2-bound mouse TPC1

EntireName: PtdIns(3,5)P2-bound mouse TPC1
Components
  • Organelle or cellular component: PtdIns(3,5)P2-bound mouse TPC1
    • Protein or peptide: Two pore calcium channel protein 1
  • Ligand: (2R)-3-{[(S)-hydroxy{[(1S,2R,3R,4S,5S,6R)-2,4,6-trihydroxy-3,5-bis(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dioctanoate
  • Ligand: 2-acetamido-2-deoxy-beta-D-glucopyranose

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Supramolecule #1: PtdIns(3,5)P2-bound mouse TPC1

SupramoleculeName: PtdIns(3,5)P2-bound mouse TPC1 / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Mus musculus (house mouse)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293F / Recombinant plasmid: pEZT-BM

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Macromolecule #1: Two pore calcium channel protein 1

MacromoleculeName: Two pore calcium channel protein 1 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 95.414758 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MAVSLDDDVP LILTLDEAES APLPPSNSLG QEQLPSKNGG SHSIHNSQVP SLVSGADSPP SSPTGHNWEM NYQEAAIYLQ EGQNNDKFF THPKDARALA AYLFVHNHFF YMMELLTALL LLLLSLCESP AVPVLKLHTY VHATLELFAL MVVVFELCMK L RWLGFHTF ...String:
MAVSLDDDVP LILTLDEAES APLPPSNSLG QEQLPSKNGG SHSIHNSQVP SLVSGADSPP SSPTGHNWEM NYQEAAIYLQ EGQNNDKFF THPKDARALA AYLFVHNHFF YMMELLTALL LLLLSLCESP AVPVLKLHTY VHATLELFAL MVVVFELCMK L RWLGFHTF VRHKRTMVKT SVLVVQFIEA IVVLVRQTSH VRVTRALRCI FLVDCRYCGG VRRNLRQIFQ SLPPFMDILL LL LFFMIIF AILGFYLFST NPSDPYFSTL ENSIVNLFVL LTTANFPDVM MPSYSRNPWS CVFFIVYLSI ELYFIMNLLL AVV FDTFND IEKHKFKSLL LHKRTAIQHA YGLLASQRRP AGISYRQFEG LMRFYKPRMS ARERFLTFKA LNQSNTPLLS LKDF YDIYE VAALQWKAKR NRQHWFDELP RTAFLIFKGI NILVNSKAFQ YFMYLVVAVN GVWILVETFM LKGGNFTSKH VPWSY LVFL TIYGVELFMK VAGLGPVEYL SSGWNLFDFS VTAFAFLGLL ALTLNMEPFY FIVVLRPLQL LRLFKLKKRY RNVLDT MFE LLPRMASLGL TLLTFYYSFA IVGMEFFNGR LTPNCCNTST VADAYRFINH TVGNKTKVEE GYYYLNNFDN ILNSFVT LF ELTVVNNWYI IMEGVTSQTS HWSRLYFMTF YIVTMVVMTI IVAFILEAFV FRMNYSRKSQ DSEVDSGIVI EKEMSKEE L MAVLELYREE RGTSSDVTRL LDTLSQMEKY QQNSMVFLGR RSRTKSDLSL KMYQEEIQEW YEEHAREQEQ QKLRGSVPG PAAQQPPGSR QRSQTVTVEA GLVPR

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Macromolecule #3: (2R)-3-{[(S)-hydroxy{[(1S,2R,3R,4S,5S,6R)-2,4,6-trihydroxy-3,5-bi...

MacromoleculeName: (2R)-3-{[(S)-hydroxy{[(1S,2R,3R,4S,5S,6R)-2,4,6-trihydroxy-3,5-bis(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dioctanoate
type: ligand / ID: 3 / Number of copies: 2 / Formula: EUJ
Molecular weightTheoretical: 746.566 Da
Chemical component information

ChemComp-EUJ:
(2R)-3-{[(S)-hydroxy{[(1S,2R,3R,4S,5S,6R)-2,4,6-trihydroxy-3,5-bis(phosphonooxy)cyclohexyl]oxy}phosphoryl]oxy}propane-1,2-diyl dioctanoate

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Macromolecule #4: 2-acetamido-2-deoxy-beta-D-glucopyranose

MacromoleculeName: 2-acetamido-2-deoxy-beta-D-glucopyranose / type: ligand / ID: 4 / Number of copies: 2 / Formula: NAG
Molecular weightTheoretical: 221.208 Da
Chemical component information

ChemComp-NAG:
2-acetamido-2-deoxy-beta-D-glucopyranose / N-Acetylglucosamine

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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

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

5e1j

Initial angle assignmentType: OTHER
Final angle assignmentType: PROJECTION MATCHING
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 82819

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