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- PDB-9p3u: Zebrafish TRPM5 E337A mutant with 5mM calcium and 0.5mM CBTA -

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Basic information

Entry
Database: PDB / ID: 9p3u
TitleZebrafish TRPM5 E337A mutant with 5mM calcium and 0.5mM CBTA
ComponentsRNA-directed RNA polymerase L,Transient receptor potential cation channel subfamily M member 5
KeywordsTRANSFERASE / TRPM5 channel / ion channel / cation channel / sodium channel / TRANSPORT PROTEIN
Function / homology
Function and homology information


NNS virus cap methyltransferase / GDP polyribonucleotidyltransferase / calcium-activated cation channel activity / bioluminescence / generation of precursor metabolites and energy / virion component / calcium channel activity / host cell cytoplasm / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / hydrolase activity ...NNS virus cap methyltransferase / GDP polyribonucleotidyltransferase / calcium-activated cation channel activity / bioluminescence / generation of precursor metabolites and energy / virion component / calcium channel activity / host cell cytoplasm / mRNA 5'-cap (guanine-N7-)-methyltransferase activity / hydrolase activity / RNA-directed RNA polymerase / RNA-directed RNA polymerase activity / calcium ion binding / ATP binding / identical protein binding / plasma membrane
Similarity search - Function
RNA-directed RNA polymerase L methyltransferase domain, rhabdovirus / Virus-capping methyltransferase, MT domain / RNA-directed RNA polymerase, paramyxovirus / Mononegavirus L protein 2-O-ribose methyltransferase / RNA-directed RNA polymerase L, C-terminal / Mononegavirus L protein 2'-O-ribose methyltransferase domain profile. / Mononegavirales RNA-directed RNA polymerase catalytic domain / Mononegavirales mRNA-capping domain V / Mononegavirales RNA dependent RNA polymerase / Mononegavirales mRNA-capping region V ...RNA-directed RNA polymerase L methyltransferase domain, rhabdovirus / Virus-capping methyltransferase, MT domain / RNA-directed RNA polymerase, paramyxovirus / Mononegavirus L protein 2-O-ribose methyltransferase / RNA-directed RNA polymerase L, C-terminal / Mononegavirus L protein 2'-O-ribose methyltransferase domain profile. / Mononegavirales RNA-directed RNA polymerase catalytic domain / Mononegavirales mRNA-capping domain V / Mononegavirales RNA dependent RNA polymerase / Mononegavirales mRNA-capping region V / RdRp of negative ssRNA viruses with non-segmented genomes catalytic domain profile. / Transient receptor potential channel, canonical / TRPM, SLOG domain / : / SLOG in TRPM / TRPM2-like domain / Green fluorescent protein, GFP / Green fluorescent protein-related / Green fluorescent protein / Green fluorescent protein / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
: / : / : / RNA-directed RNA polymerase L / Transient receptor potential cation channel subfamily M member 5
Similarity search - Component
Biological speciesDanio rerio (zebrafish)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.75 Å
AuthorsRuan, Z. / Du, J. / Lu, W.
Funding support United States, 6items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R00NS128258 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)R01HL153219 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01NS112363 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM138321 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01NS111031 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01NS129804 United States
CitationJournal: Nat Chem Biol / Year: 2026
Title: A single allosteric site merges activation, modulation and inhibition in TRPM5.
Authors: Zheng Ruan / Junuk Lee / Yangyang Li / Ian J Orozco / Juan Du / Wei Lü /
Abstract: TRPM5 is a Ca-activated monovalent cation channel essential for taste perception, insulin secretion and gastrointestinal chemosensation. Canonical TRPM5 activation requires Ca binding at two distinct ...TRPM5 is a Ca-activated monovalent cation channel essential for taste perception, insulin secretion and gastrointestinal chemosensation. Canonical TRPM5 activation requires Ca binding at two distinct sites: an agonist site within the lower vestibule of the S1-S4 pocket in the transmembrane domain (Ca) and a modulatory site in the intracellular domain (Ca) that tunes voltage dependence and agonist sensitivity. Here we characterize CBTA as a noncalcium agonist that binds to the upper vestibule of the S1-S4 pocket, directly above Ca. CBTA alone mimics the dual role of Ca and Ca, merging agonist activation with voltage modulation. CBTA also renders TRPM5 supersensitive to Ca, synergistically hyperactivating the channel even at near-resting Ca levels. We further demonstrate that the inhibitor triphenylphosphine oxide binds the same site but stabilizes a nonconductive state. These opposing effects reveal the upper S1-S4 pocket as a multifunctional regulatory hub integrating activation, inhibition and modulation in TRPM5.
History
DepositionJun 14, 2025Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 7, 2026Provider: repository / Type: Initial release
Revision 1.0Jan 7, 2026Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
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Revision 1.0Jan 7, 2026Data content type: FSC / Data content type: FSC / Provider: repository / Type: Initial release
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Revision 1.1Jan 21, 2026Group: Data collection / Database references ...Data collection / Database references / Derived calculations / Structure summary
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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
C: RNA-directed RNA polymerase L,Transient receptor potential cation channel subfamily M member 5
A: RNA-directed RNA polymerase L,Transient receptor potential cation channel subfamily M member 5
B: RNA-directed RNA polymerase L,Transient receptor potential cation channel subfamily M member 5
D: RNA-directed RNA polymerase L,Transient receptor potential cation channel subfamily M member 5
hetero molecules


Theoretical massNumber of molelcules
Total (without water)674,37924
Polymers667,0314
Non-polymers7,34920
Water1448
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 / Sugars , 2 types, 8 molecules CABD

#1: Protein
RNA-directed RNA polymerase L,Transient receptor potential cation channel subfamily M member 5 / Large structural protein / Replicase / Transcriptase


Mass: 166757.641 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Danio rerio (zebrafish) / Gene: trpm5 / Production host: Homo sapiens (human)
References: UniProt: A0A5P9VSM8, UniProt: S5UH55, NNS virus cap methyltransferase, RNA-directed RNA polymerase, GDP polyribonucleotidyltransferase
#2: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C8H15NO6 / Feature type: SUBJECT OF INVESTIGATION
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 5 types, 24 molecules

#3: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Ca / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical
ChemComp-A1CGZ / Tigogenin


Mass: 416.636 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C27H44O3
#5: Chemical
ChemComp-A1CGW / 5-chloro-N-[(5-chloro-1,3-thiazol-2-yl)methyl]-1,2-benzothiazol-6-amine


Mass: 316.229 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: C11H7Cl2N3S2 / Feature type: SUBJECT OF INVESTIGATION
#6: Chemical
ChemComp-A1CG0 / (2R)-2-(hydroxymethyl)-4-{[(10xi,13xi,22xi,25R)-5beta,14beta,17beta-spirostan-3beta-yl]oxy}butyl 4-O-alpha-D-glucopyranosyl-beta-D-glucopyranoside


Mass: 843.049 Da / Num. of mol.: 4 / Source method: isolated from a natural source / Formula: C44H74O15
#7: Water ChemComp-HOH / water


Mass: 18.015 Da / Num. of mol.: 8 / Source method: isolated from a natural source / Formula: H2O

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Details

Has ligand of interestY
Has protein modificationY

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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: zebrafish TRPM5 E337A mutant in GDN detergent with 5mM calcium and 0.5mM CBTA
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Danio rerio (zebrafish)
Source (recombinant)Organism: Homo sapiens (human)
Buffer solutionpH: 8
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: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1900 nm / Nominal defocus min: 1100 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1cryoSPARC4particle selection
13RELION53D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.75 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 192713 / Symmetry type: POINT

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