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- PDB-8sla: Cryo-EM structure of the rat TRPM5 channel in trace calcium, trace-2 -

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Basic information

Entry
Database: PDB / ID: 8sla
TitleCryo-EM structure of the rat TRPM5 channel in trace calcium, trace-2
ComponentsTransient receptor potential cation channel subfamily M member 5
KeywordsMEMBRANE PROTEIN / ion channel / Transient Receptor Potential / TRP / Transient Receptor Potential Melastatin 5 / TRPM5
Function / homology
Function and homology information


metal ion transport / calcium-activated cation channel activity / sodium channel activity / monoatomic cation transmembrane transport / monoatomic ion channel activity / potassium channel activity / positive regulation of insulin secretion involved in cellular response to glucose stimulus / monoatomic ion transmembrane transport / neuronal cell body / dendrite / plasma membrane
Similarity search - Function
TRPM, SLOG domain / : / SLOG in TRPM / TRPM2-like domain / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Transient receptor potential cation channel subfamily M member 5
Similarity search - Component
Biological speciesRattus norvegicus (Norway rat)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsKaruppan, S. / Schrag, L.G. / Jara-Oseguera, A. / Zubcevic, L.
Funding support United States, 2items
OrganizationGrant numberCountry
Other private United States
Other privaten/a, Start-up funds
CitationJournal: Proc Natl Acad Sci U S A / Year: 2024
Title: Structural dynamics at cytosolic interprotomer interfaces control gating of a mammalian TRPM5 channel.
Authors: Sebastian Karuppan / Lynn Goss Schrag / Caroline M Pastrano / Andrés Jara-Oseguera / Lejla Zubcevic /
Abstract: The transient receptor potential melastatin (TRPM) tetrameric cation channels are involved in a wide range of biological functions, from temperature sensing and taste transduction to regulation of ...The transient receptor potential melastatin (TRPM) tetrameric cation channels are involved in a wide range of biological functions, from temperature sensing and taste transduction to regulation of cardiac function, inflammatory pain, and insulin secretion. The structurally conserved TRPM cytoplasmic domains make up >70 % of the total protein. To investigate the mechanism by which the TRPM cytoplasmic domains contribute to gating, we employed electrophysiology and cryo-EM to study TRPM5-a channel that primarily relies on activation via intracellular Ca. Here, we show that activation of mammalian TRPM5 channels is strongly altered by Ca-dependent desensitization. Structures of rat TRPM5 identify a series of conformational transitions triggered by Ca binding, whereby formation and dissolution of cytoplasmic interprotomer interfaces appear to control activation and desensitization of the channel. This study shows the importance of the cytoplasmic assembly in TRPM5 channel function and sets the stage for future investigations of other members of the TRPM family.
History
DepositionApr 21, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 3, 2024Provider: repository / Type: Initial release
Revision 1.1Jul 10, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.name / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
D: Transient receptor potential cation channel subfamily M member 5
C: Transient receptor potential cation channel subfamily M member 5
A: Transient receptor potential cation channel subfamily M member 5
B: Transient receptor potential cation channel subfamily M member 5


Theoretical massNumber of molelcules
Total (without water)529,4074
Polymers529,4074
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Transient receptor potential cation channel subfamily M member 5


Mass: 132351.719 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Rattus norvegicus (Norway rat) / Gene: Trpm5 / Cell line (production host): HEK293S GnTl- / Production host: Homo sapiens (human) / References: UniProt: A0A455XI77
Has ligand of interestN

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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 Melastatin 5 (TRPM5) / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.52 MDa / Experimental value: NO
Source (natural)Organism: Rattus norvegicus (Norway rat)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK 293S GnTl- / Plasmid: pEGBacMam
Buffer solutionpH: 8
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: LEICA EM GP / Cryogen name: ETHANE / Humidity: 85 % / Chamber temperature: 296.15 K

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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: OTHER
Electron lensMode: BRIGHT FIELD / Nominal magnification: 81000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 45 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV

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Processing

EM software
IDNameVersionCategory
7Cootmodel fitting
9PHENIXmodel refinement
13cryoSPARC3.3.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 2601822
SymmetryPoint symmetry: C4 (4 fold cyclic)
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 139580 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT
Atomic model buildingAccession code: 8SL6 / Details: model generated in this study / Source name: Other / Type: other

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