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- EMDB-20482: Human TRPM2 bound to 8-Br-cADPR and calcium -

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Basic information

Entry
Database: EMDB / ID: EMD-20482
TitleHuman TRPM2 bound to 8-Br-cADPR and calcium
Map datahuman TRPM2 bound with 8-Br-cADPR and calcium, unsharpened map
Sample
  • Complex: human TRPM2
    • Protein or peptide: Transient receptor potential cation channel subfamily M member 2
  • Ligand: (2R,3R,4S,5R,13R,14S,15R,16R)-24-amino-18-bromo-3,4,14,15-tetrahydroxy-7,9,11,25,26-pentaoxa-17,19,22-triaza-1-azonia-8,10-diphosphapentacyclo[18.3.1.1^2,5^.1^13,16^.0^17,21^]hexacosa-1(24),18,20,22-tetraene-8,10-diolate 8,10-dioxide
  • Ligand: CALCIUM IONCalcium
Function / homology
Function and homology information


cellular response to purine-containing compound / mono-ADP-D-ribose binding / manganese ion transmembrane transporter activity / zinc ion transmembrane transport / dendritic cell differentiation / ligand-gated calcium channel activity / response to purine-containing compound / cellular response to temperature stimulus / regulation of filopodium assembly / sodium channel activity ...cellular response to purine-containing compound / mono-ADP-D-ribose binding / manganese ion transmembrane transporter activity / zinc ion transmembrane transport / dendritic cell differentiation / ligand-gated calcium channel activity / response to purine-containing compound / cellular response to temperature stimulus / regulation of filopodium assembly / sodium channel activity / regulation of actin cytoskeleton reorganization / TRP channels / response to hydroperoxide / dendritic cell chemotaxis / calcium ion transmembrane import into cytosol / : / estrous cycle / temperature homeostasis / calcium-release channel activity / calcium ion import across plasma membrane / ficolin-1-rich granule membrane / tertiary granule membrane / calcium-mediated signaling using intracellular calcium source / specific granule membrane / release of sequestered calcium ion into cytosol / monoatomic cation channel activity / cellular response to calcium ion / calcium ion transmembrane transport / calcium channel activity / cytoplasmic vesicle membrane / positive regulation of insulin secretion / cellular response to hydrogen peroxide / calcium ion transport / perikaryon / protein homotetramerization / response to heat / lysosomal membrane / lysosome / neuron projection / Neutrophil degranulation / calcium ion binding / plasma membrane
Similarity search - Function
TRPM, SLOG domain / SLOG in TRPM / NUDIX hydrolase domain / Nudix hydrolase domain profile. / NUDIX hydrolase-like domain superfamily / Ion transport domain / Ion transport protein
Similarity search - Domain/homology
Transient receptor potential cation channel subfamily M member 2
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsDu J / Lu W / Huang Y
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)R01NS111031 United States
CitationJournal: Elife / Year: 2019
Title: Ligand recognition and gating mechanism through three ligand-binding sites of human TRPM2 channel.
Authors: Yihe Huang / Becca Roth / Wei Lü / Juan Du /
Abstract: TRPM2 is critically involved in diverse physiological processes including core temperature sensing, apoptosis, and immune response. TRPM2's activation by Ca and ADP ribose (ADPR), an NAD-metabolite ...TRPM2 is critically involved in diverse physiological processes including core temperature sensing, apoptosis, and immune response. TRPM2's activation by Ca and ADP ribose (ADPR), an NAD-metabolite produced under oxidative stress and neurodegenerative conditions, suggests a role in neurological disorders. We provide a central concept between triple-site ligand binding and the channel gating of human TRPM2. We show consecutive structural rearrangements and channel activation of TRPM2 induced by binding of ADPR in two indispensable locations, and the binding of Ca in the transmembrane domain. The 8-Br-cADPR-an antagonist of cADPR-binds only to the MHR1/2 domain and inhibits TRPM2 by stabilizing the channel in an apo-like conformation. We conclude that MHR1/2 acts as a orthostatic ligand-binding site for TRPM2. The NUDT9-H domain binds to a second ADPR to assist channel activation in vertebrates, but not necessary in invertebrates. Our work provides insights into the gating mechanism of human TRPM2 and its pharmacology.
History
DepositionJul 18, 2019-
Header (metadata) releaseJul 31, 2019-
Map releaseSep 25, 2019-
UpdateDec 18, 2019-
Current statusDec 18, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.009
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.009
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6puu
  • Surface level: 0.009
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20482.png

Downloads & links

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Map

FileDownload / File: emd_20482.map.gz / Format: CCP4 / Size: 103 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationhuman TRPM2 bound with 8-Br-cADPR and calcium, unsharpened map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 300 pix.
= 322.2 Å		1.07 Å/pix.
x 300 pix.
= 322.2 Å		1.07 Å/pix.
x 300 pix.
= 322.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.074 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.009 / Movie #1: 0.009
Minimum - Maximum-0.0210351 - 0.04778512
Average (Standard dev.)0.00004284061 (±0.0024705718)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions300300300
Spacing300300300
CellA=B=C: 322.2 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0741.0741.074
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z322.200322.200322.200
α/β/γ90.00090.00090.000
start NX/NY/NZ1548552
NX/NY/NZ198170217
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS300300300
D min/max/mean-0.0210.0480.000

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Supplemental data

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Sample components

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Entire : human TRPM2

EntireName: human TRPM2
Components
  • Complex: human TRPM2
    • Protein or peptide: Transient receptor potential cation channel subfamily M member 2
  • Ligand: (2R,3R,4S,5R,13R,14S,15R,16R)-24-amino-18-bromo-3,4,14,15-tetrahydroxy-7,9,11,25,26-pentaoxa-17,19,22-triaza-1-azonia-8,10-diphosphapentacyclo[18.3.1.1^2,5^.1^13,16^.0^17,21^]hexacosa-1(24),18,20,22-tetraene-8,10-diolate 8,10-dioxide
  • Ligand: CALCIUM IONCalcium

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Supramolecule #1: human TRPM2

SupramoleculeName: human TRPM2 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human)

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Macromolecule #1: Transient receptor potential cation channel subfamily M member 2

MacromoleculeName: Transient receptor potential cation channel subfamily M member 2
type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 172.280062 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:

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Macromolecule #2: (2R,3R,4S,5R,13R,14S,15R,16R)-24-amino-18-bromo-3,4,14,15-tetrahy...

MacromoleculeName: (2R,3R,4S,5R,13R,14S,15R,16R)-24-amino-18-bromo-3,4,14,15-tetrahydroxy-7,9,11,25,26-pentaoxa-17,19,22-triaza-1-azonia-8,10-diphosphapentacyclo[18.3.1.1^2,5^.1^13,16^.0^17,21^]hexacosa-1(24) ...Name: (2R,3R,4S,5R,13R,14S,15R,16R)-24-amino-18-bromo-3,4,14,15-tetrahydroxy-7,9,11,25,26-pentaoxa-17,19,22-triaza-1-azonia-8,10-diphosphapentacyclo[18.3.1.1^2,5^.1^13,16^.0^17,21^]hexacosa-1(24),18,20,22-tetraene-8,10-diolate 8,10-dioxide
type: ligand / ID: 2 / Number of copies: 4 / Formula: CV1
Molecular weightTheoretical: 621.204 Da

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Macromolecule #3: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 3 / Number of copies: 4 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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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
GridDetails: unspecified
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 1-40 / Average exposure time: 8.0 sec. / Average electron dose: 6.8 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware - Name: Gctf
Startup modelType of model: OTHER / Details: cryosparc
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3) / Number images used: 102259

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-6puu:
Human TRPM2 bound to 8-Br-cADPR and calcium

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