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- EMDB-40867: NUDT9-H domain focused cryo-EM map of TRPM2 chanzyme in the prese... -

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Basic information

Entry
Database: EMDB / ID: EMD-40867
TitleNUDT9-H domain focused cryo-EM map of TRPM2 chanzyme in the presence of Magnesium, Adenosine monophosphate, and Ribose-5-phosphate
Map data
Sample
  • Complex: TRPM2 chanzyme incubated with Magnesium and ADP-ribose for 4min, ADP-ribose completely hydrolyzed to Adenosine monophosphate, and Ribose-5-phosphate
    • Protein or peptide: TRPM2 chanzyme
KeywordsTRPM2 Chanzyme / Channel-enzyme / MEMBRANE PROTEIN
Biological speciesSalpingoeca rosetta (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.83 Å
AuthorsHuang Y / Sushant K / Lu W / Du J
Funding support United States, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL153219 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS112363 United States
National Institutes of Health/National Institute of Neurological Disorders and Stroke (NIH/NINDS)NS111031 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM129547 United States
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Coupling enzymatic activity and gating in an ancient TRPM chanzyme and its molecular evolution.
Authors: Yihe Huang / Sushant Kumar / Junuk Lee / Wei Lü / Juan Du /
Abstract: Channel enzymes represent a class of ion channels with enzymatic activity directly or indirectly linked to their channel function. We investigated a TRPM2 chanzyme from choanoflagellates that ...Channel enzymes represent a class of ion channels with enzymatic activity directly or indirectly linked to their channel function. We investigated a TRPM2 chanzyme from choanoflagellates that integrates two seemingly incompatible functions into a single peptide: a channel module activated by ADP-ribose with high open probability and an enzyme module (NUDT9-H domain) consuming ADP-ribose at a remarkably slow rate. Using time-resolved cryogenic-electron microscopy, we captured a complete series of structural snapshots of gating and catalytic cycles, revealing the coupling mechanism between channel gating and enzymatic activity. The slow kinetics of the NUDT9-H enzyme module confers a self-regulatory mechanism: ADPR binding triggers NUDT9-H tetramerization, promoting channel opening, while subsequent hydrolysis reduces local ADPR, inducing channel closure. We further demonstrated how the NUDT9-H domain has evolved from a structurally semi-independent ADP-ribose hydrolase module in early species to a fully integrated component of a gating ring essential for channel activation in advanced species.
History
DepositionMay 25, 2023-
Header (metadata) releaseMay 29, 2024-
Map releaseMay 29, 2024-
UpdateOct 30, 2024-
Current statusOct 30, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40867.png

Downloads & links

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Map

FileDownload / File: emd_40867.map.gz / Format: CCP4 / Size: 325 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 440 pix.
= 363.44 Å		0.83 Å/pix.
x 440 pix.
= 363.44 Å		0.83 Å/pix.
x 440 pix.
= 363.44 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.826 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.008
Minimum - Maximum-0.0039436226 - 0.027939118
Average (Standard dev.)-0.00016799387 (±0.0005258301)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions440440440
Spacing440440440
CellA=B=C: 363.44 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #1

Fileemd_40867_additional_1.map
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Half map: #1

Fileemd_40867_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_40867_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : TRPM2 chanzyme incubated with Magnesium and ADP-ribose for 4min, ...

EntireName: TRPM2 chanzyme incubated with Magnesium and ADP-ribose for 4min, ADP-ribose completely hydrolyzed to Adenosine monophosphate, and Ribose-5-phosphate
Components
  • Complex: TRPM2 chanzyme incubated with Magnesium and ADP-ribose for 4min, ADP-ribose completely hydrolyzed to Adenosine monophosphate, and Ribose-5-phosphate
    • Protein or peptide: TRPM2 chanzyme

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Supramolecule #1: TRPM2 chanzyme incubated with Magnesium and ADP-ribose for 4min, ...

SupramoleculeName: TRPM2 chanzyme incubated with Magnesium and ADP-ribose for 4min, ADP-ribose completely hydrolyzed to Adenosine monophosphate, and Ribose-5-phosphate
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Salpingoeca rosetta (eukaryote)

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Macromolecule #1: TRPM2 chanzyme

MacromoleculeName: TRPM2 chanzyme / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Salpingoeca rosetta (eukaryote)
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: NRL NSAVAV HGHTAEAAEW YVPPEEYPKS GGVKRYLIDA SMVPLSIMCP SYDPVEYTHP SVAAQPVWAD PADPRKIKFN VKDE VNGKV VDRTSCHPSG ISIDSNTGRP INPWGRTGMT GRGLLGKWGV NQAADTVVTR WKRSPDGSIL ERDGKKVLEF VAIQR QDNK ...String:
NRL NSAVAV HGHTAEAAEW YVPPEEYPKS GGVKRYLIDA SMVPLSIMCP SYDPVEYTHP SVAAQPVWAD PADPRKIKFN VKDE VNGKV VDRTSCHPSG ISIDSNTGRP INPWGRTGMT GRGLLGKWGV NQAADTVVTR WKRSPDGSIL ERDGKKVLEF VAIQR QDNK MWAIPGGFVD NGEDVALTSG REFMEEALGM GTSADLMSAE SKDSLAALFS SGTIVARIYC EDPRNTDNAW VETTCV NFH DESGRHAARL KLQGGDDAEH ARWMMVHGGL NLFASHRTLL QHVTSALNAY F(RP5)(AMP)(MG)(MG)

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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

Concentration8.5 mg/mL
BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average exposure time: 0.02 sec. / Average electron dose: 49.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.9000000000000001 µm / Nominal defocus min: 0.9 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.83 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 634404
Initial angle assignmentType: RANDOM ASSIGNMENT
Final angle assignmentType: MAXIMUM LIKELIHOOD

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL

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