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- PDB-5w3s: Cryo-electron microscopy structure of a TRPML3 ion channel -

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Basic information

Entry
Database: PDB / ID: 5w3s
TitleCryo-electron microscopy structure of a TRPML3 ion channel
ComponentsMucolipin-3 isoform 1
KeywordsTRANSPORT PROTEIN / transient receptor potential channel / mucolipin / ion channel / membrane transport / TRPML / TRP channel / calcium channel / PIP2 / PI(3 / 5)P2 / lipid-gated channel / mucolipidosis / lysosomal ion channel / lysosome
Function / homology
Function and homology information


stereocilium membrane / : / intracellularly phosphatidylinositol-3,5-bisphosphate-gated monatomic cation channel activity / inner ear auditory receptor cell differentiation / phosphatidylinositol-3,5-bisphosphate binding / monoatomic cation transmembrane transport / autophagosome membrane / locomotory behavior / late endosome membrane / early endosome membrane ...stereocilium membrane / : / intracellularly phosphatidylinositol-3,5-bisphosphate-gated monatomic cation channel activity / inner ear auditory receptor cell differentiation / phosphatidylinositol-3,5-bisphosphate binding / monoatomic cation transmembrane transport / autophagosome membrane / locomotory behavior / late endosome membrane / early endosome membrane / protein homotetramerization / membrane => GO:0016020
Similarity search - Function
Mucolipin / Polycystin cation channel, PKD1/PKD2 / Polycystin cation channel
Similarity search - Domain/homology
1,2-Distearoyl-sn-glycerophosphoethanolamine / CHOLESTEROL HEMISUCCINATE / Mucolipin-3
Similarity search - Component
Biological speciesCallithrix jacchus (white-tufted-ear marmoset)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.94 Å
AuthorsHirschi, M. / Herzik, M.A. / Wie, J. / Suo, Y. / Borschel, W.F. / Ren, D. / Lander, G.C. / Lee, S.Y.
Funding support United States, 2items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R35NS097241 United States
National Institutes of Health/National Institute of Biomedical Imaging and Bioengineering (NIH/NIBIB)DP2EB020402 United States
CitationJournal: Nature / Year: 2017
Title: Cryo-electron microscopy structure of the lysosomal calcium-permeable channel TRPML3.
Authors: Marscha Hirschi / Mark A Herzik / Jinhong Wie / Yang Suo / William F Borschel / Dejian Ren / Gabriel C Lander / Seok-Yong Lee /
Abstract: The modulation of ion channel activity by lipids is increasingly recognized as a fundamental component of cellular signalling. The transient receptor potential mucolipin (TRPML) channel family ...The modulation of ion channel activity by lipids is increasingly recognized as a fundamental component of cellular signalling. The transient receptor potential mucolipin (TRPML) channel family belongs to the TRP superfamily and is composed of three members: TRPML1-TRPML3. TRPMLs are the major Ca-permeable channels on late endosomes and lysosomes (LEL). They regulate the release of Ca from organelles, which is important for various physiological processes, including organelle trafficking and fusion. Loss-of-function mutations in the MCOLN1 gene, which encodes TRPML1, cause the neurodegenerative lysosomal storage disorder mucolipidosis type IV, and a gain-of-function mutation (Ala419Pro) in TRPML3 gives rise to the varitint-waddler (Va) mouse phenotype. Notably, TRPML channels are activated by the low-abundance and LEL-enriched signalling lipid phosphatidylinositol-3,5-bisphosphate (PtdIns(3,5)P), whereas other phosphoinositides such as PtdIns(4,5)P, which is enriched in plasma membranes, inhibit TRPMLs. Conserved basic residues at the N terminus of the channel are important for activation by PtdIns(3,5)P and inhibition by PtdIns(4,5)P. However, owing to a lack of structural information, the mechanism by which TRPML channels recognize PtdIns(3,5)P and increase their Ca conductance remains unclear. Here we present the cryo-electron microscopy (cryo-EM) structure of a full-length TRPML3 channel from the common marmoset (Callithrix jacchus) at an overall resolution of 2.9 Å. Our structure reveals not only the molecular basis of ion conduction but also the unique architecture of TRPMLs, wherein the voltage sensor-like domain is linked to the pore via a cytosolic domain that we term the mucolipin domain. Combined with functional studies, these data suggest that the mucolipin domain is responsible for PtdIns(3,5)P binding and subsequent channel activation, and that it acts as a 'gating pulley' for lipid-dependent TRPML gating.
History
DepositionJun 8, 2017Deposition site: RCSB / Processing site: RCSB
Revision 1.0Oct 11, 2017Provider: repository / Type: Initial release
Revision 1.1Oct 25, 2017Group: Database references / Category: citation / Item: _citation.pdbx_database_id_PubMed / _citation.title
Revision 1.2Nov 1, 2017Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.3Dec 11, 2019Group: Author supporting evidence / Category: pdbx_audit_support / Item: _pdbx_audit_support.funding_organization
Revision 1.4Mar 13, 2024Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Structure summary
Category: chem_comp / chem_comp_atom ...chem_comp / chem_comp_atom / chem_comp_bond / database_2 / entity / pdbx_entity_nonpoly
Item: _chem_comp.name / _chem_comp.pdbx_synonyms ..._chem_comp.name / _chem_comp.pdbx_synonyms / _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _entity.pdbx_description / _pdbx_entity_nonpoly.name

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Structure visualization

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Assembly

Deposited unit
A: Mucolipin-3 isoform 1
B: Mucolipin-3 isoform 1
C: Mucolipin-3 isoform 1
D: Mucolipin-3 isoform 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)269,34026
Polymers260,3694
Non-polymers8,97122
Water724
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area33250 Å2
ΔGint-356 kcal/mol
Surface area87220 Å2
MethodPISA

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Components

#1: Protein
Mucolipin-3 isoform 1


Mass: 65092.301 Da / Num. of mol.: 4 / Mutation: N138Q
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Callithrix jacchus (white-tufted-ear marmoset)
Gene: MCOLN3 / Plasmid: pFastBac / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: F6RG56
#2: Chemical
ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 12 / Source method: obtained synthetically / Formula: C31H50O4
#3: Chemical
ChemComp-NA / SODIUM ION


Mass: 22.990 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Na
#4: Chemical
ChemComp-3PE / 1,2-Distearoyl-sn-glycerophosphoethanolamine / 3-SN-PHOSPHATIDYLETHANOLAMINE / 1,2-DIACYL-SN-GLYCERO-3-PHOSPHOETHANOLAMINE / Phosphatidylethanolamine


Mass: 748.065 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C41H82NO8P / Comment: phospholipid*YM
#5: Water ChemComp-HOH / water / Water


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

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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 Mucolipin 3 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.26 MDa / Experimental value: NO
Source (natural)Organism: Callithrix jacchus (white-tufted-ear marmoset)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm) / Strain: Sf9 / Plasmid: pFastBac
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 FIELDBright-field microscopy
Image recordingAverage exposure time: 12 sec. / Electron dose: 63 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.11.1_2575: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 2.94 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 104084 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00631924
ELECTRON MICROSCOPYf_angle_d1.37557196
ELECTRON MICROSCOPYf_dihedral_angle_d9.38413092
ELECTRON MICROSCOPYf_chiral_restr0.052588
ELECTRON MICROSCOPYf_plane_restr0.0044784

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