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- EMDB-20367: Cryo-EM structure of the zebrafish TRPM2 channel in the apo confo... -

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Basic information

Entry
Database: EMDB / ID: EMD-20367
TitleCryo-EM structure of the zebrafish TRPM2 channel in the apo conformation, processed with C4 symmetry
Map datazebrafish TRPM2 channel in the apo conformation, processed with C4 symmetry
Sample
  • Complex: Transient receptor potential cation channel subfamily M member 2
    • Protein or peptide: Transient receptor potential cation channel subfamily M member 2
Keywordswarmth sensor / redox sensor / calcium-permeable ion channel / TRP channel / TRPM channel / ion channel / ADP-ribose / TRANSPORT PROTEIN
Function / homology
Function and homology information


TRP channels / Neutrophil degranulation / ADP-D-ribose binding / mono-ADP-D-ribose binding / ligand-gated calcium channel activity / ligand-gated monoatomic cation channel activity / monoatomic ion channel activity / calcium channel activity / calcium ion transmembrane transport / protein homotetramerization ...TRP channels / Neutrophil degranulation / ADP-D-ribose binding / mono-ADP-D-ribose binding / ligand-gated calcium channel activity / ligand-gated monoatomic cation channel activity / monoatomic ion channel activity / calcium channel activity / calcium ion transmembrane transport / protein homotetramerization / calcium ion binding / plasma membrane
Similarity search - Function
TRPM, SLOG domain / : / SLOG in TRPM / 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 speciesDanio rerio (zebrafish)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.3 Å
AuthorsYin Y / Wu M
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS) United States
CitationJournal: Nat Commun / Year: 2019
Title: Visualizing structural transitions of ligand-dependent gating of the TRPM2 channel.
Authors: Ying Yin / Mengyu Wu / Allen L Hsu / William F Borschel / Mario J Borgnia / Gabriel C Lander / Seok-Yong Lee /
Abstract: The transient receptor potential melastatin 2 (TRPM2) channel plays a key role in redox sensation in many cell types. Channel activation requires binding of both ADP-ribose (ADPR) and Ca. The ...The transient receptor potential melastatin 2 (TRPM2) channel plays a key role in redox sensation in many cell types. Channel activation requires binding of both ADP-ribose (ADPR) and Ca. The recently published TRPM2 structures from Danio rerio in the ligand-free and the ADPR/Ca-bound conditions represent the channel in closed and open states, which uncovered substantial tertiary and quaternary conformational rearrangements. However, it is unclear how these rearrangements are achieved within the tetrameric channel during channel gating. Here we report the cryo-electron microscopy structures of Danio rerio TRPM2 in the absence of ligands, in complex with Ca alone, and with both ADPR and Ca, resolved to ~4.3 Å, ~3.8 Å, and ~4.2 Å, respectively. In contrast to the published results, our studies capture ligand-bound TRPM2 structures in two-fold symmetric intermediate states, offering a glimpse of the structural transitions that bridge the closed and open conformations.
History
DepositionJun 30, 2019-
Header (metadata) releaseAug 28, 2019-
Map releaseAug 28, 2019-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6pkv
  • Surface level: 0.01
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20367.png

Downloads & links

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Map

FileDownload / File: emd_20367.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationzebrafish TRPM2 channel in the apo conformation, processed with C4 symmetry
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.07 Å/pix.
x 256 pix.
= 272.896 Å		1.07 Å/pix.
x 256 pix.
= 272.896 Å		1.07 Å/pix.
x 256 pix.
= 272.896 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.066 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum-0.027883323 - 0.055690564
Average (Standard dev.)-0.000032200187 (±0.0030542659)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin0-2560
Dimensions256256256
Spacing256256256
CellA=B=C: 272.896 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0661.0661.066
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z272.896272.896272.896
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ167149147
MAP C/R/S123
start NC/NR/NS-25600
NC/NR/NS256256256
D min/max/mean-0.0280.056-0.000

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Supplemental data

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Sample components

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Entire : Transient receptor potential cation channel subfamily M member 2

EntireName: Transient receptor potential cation channel subfamily M member 2
Components
  • Complex: Transient receptor potential cation channel subfamily M member 2
    • Protein or peptide: Transient receptor potential cation channel subfamily M member 2

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

SupramoleculeName: Transient receptor potential cation channel subfamily M member 2
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Danio rerio (zebrafish)

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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: Danio rerio (zebrafish)
Molecular weightTheoretical: 167.608625 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MALGTSGVKI HPNGNSNQLG VQLENVKLTS LFKKLDKRCS LASWIKENIK KKECCFYVED GREGICKCGY PKVQHCDEAI KPEDYMGEQ WDKHRHVRET PTDAFGDISF GGLGQKTGKY VRVSSDTSCE NLYQLMTEQW KLRSPNLLIS VTGGAKNFYI K THLKDKFR ...String:
MALGTSGVKI HPNGNSNQLG VQLENVKLTS LFKKLDKRCS LASWIKENIK KKECCFYVED GREGICKCGY PKVQHCDEAI KPEDYMGEQ WDKHRHVRET PTDAFGDISF GGLGQKTGKY VRVSSDTSCE NLYQLMTEQW KLRSPNLLIS VTGGAKNFYI K THLKDKFR RGLIKVAQTT GAWILTGGTH AGVMKHVGMA VRDYTLSSGS MEGQIVVIGV APWGVIHNRS TLIHPEGRFP AY YSLDEQG QGRLSCLDIN HTHFLLVDDG TQGHYGVEIE LRARLEKLIS KLSLGNRESG VTIPVVCVVL DGGPGTLNTI YNS MLNHTP CVVLEGSGRL ADVIAHVASV PVSKVTMALI NRLLKRFFMQ EYKNFTELQI IEWTKKIQDI LRMPHLLTVF RIDE DKNYD VDVAILQALL KASRSDEHAG RHCWERQLEL AVAWNRVDIA ESEIFTEESQ WTSSDLHPAM FSALVGDKPE FVRLL LENG VCVREFLERE ETLCELYSHL PSCFFLRKLA KRVQGGKMRR GQEPLPGSRK VCLSHVSEEV RHLLGSFTQP LYIASR YKP TKDDVRLKVP SKGALDLPCS GEEWSADTVW DPGRDLFLWA VVQNNRELAE IGWEQCRDCI AAALAASKIL RKLAQES GE DDSEEATEML ELANHYEKQA IGVFSECHSW DAQRAQKLLI RISPSWGRST CLWLALEAHD KSFIAHSGVQ ALLTQIWC G ELSVDNPHWK VLLCMIFFPL IYTGFLTFRR DEDIQRQAER TEQQKLAMES VFAGQSDGKI KRHLRGFSQK SELKPLNCS SRLMSFLKSP QVKFYWNIAS YFGFLWLFAV VLMIDFQTSP SWRELLLYVW LTSLVCEEIR QLYHDFDGSG FRRKAKMYIK DLWNILDVL SIVLFIAGLI CRLQASDTVF YIGKVILCID FIIFCLRLMA IFSISRTLGP KIIIVRRMML DLFFFMFLLS I WVVAYGVA KQGILIENEE RLNWIIRGAV YEPYITIFGN FPTNIDNTLF DISSCSVNAS DPLKPKCPML NADNTPVFPE WL TIMMLCV YLLFANILLL NLLIAIFNYT FQEVQDNTDT IWKFQRYELI KEYHSRPALP PPFILLSHLI LFIRGVFLRD LPQ RHKNFR QELEQTEEEE LLSWEAYMKD NYLASTRQDE SQSVEHRIHD TAEKVGAMSE LLEREQEMV(UNK) (UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK)(UNK) (UNK)(UNK)(UNK)DEEAPHM FARQLQYPDS TVRRFPVPEE KVSWEVNFSP YQPPVYNQQD SSESDTSALD KHRNPGGR T GIRGKGALNT LGPNHILHPI FTRWRDAEHK VLEFLAVWED AEKRWALLGG PAQPDEPLAQ VLERILGKKL NEKTKTLLK AGEEVYKGYV DDSRNTDNAW VETSIITLHC DKNTPLMADL NHMVESSLSS HQPLQWREVS SDACRCSYQR EALRQIAHHH NTYFSNSLE VLFQGPDYKD DDDKAHHHHH HHHHH

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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

Concentration3.6 mg/mL
BufferpH: 8
GridModel: UltrAuFoil / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Details: 15 mA
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 298.15 K / Instrument: LEICA EM GP

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 3776 / Average exposure time: 4.6 sec. / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.75 µm / Nominal magnification: 22500
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 301199
Startup modelType of model: EMDB MAP
EMDB ID:

7127


Details: The EM map for the previously reported apo TRPM8 structure (EMD-7127) was low-pass filtered to 30 Angstrom and used as an initial model without a reference mask.
Final reconstructionApplied symmetry - Point group: C4 (4 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0)
Details: No symmetry (C1) was imposed during reconstructions and classifications. C4 symmetry was imposed during the final 3D auto-refinement.
Number images used: 58005
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 3.0)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 3.0)

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Atomic model buiding 1

Initial modelPDB ID:

6d73


Chain - Chain ID: A / Chain - Source name: PDB / Chain - Initial model type: experimental model
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 150
Output model

PDB-6pkv:
Cryo-EM structure of the zebrafish TRPM2 channel in the apo conformation, processed with C4 symmetry

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