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- EMDB-41458: nhTMEM16 lipid scramblase in lipid nanodiscs with MSP1E3 scaffold... -

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Basic information

Entry
Database: EMDB / ID: EMD-41458
TitlenhTMEM16 lipid scramblase in lipid nanodiscs with MSP1E3 scaffold protein in the presence of Ca2+ (closed state)(local refined monomer map)
Map dataLocal refined monomer map used to construct the composite map of nhTMEM16 in lipid nanodiscs with MSP1E3 scaffold protein in the presence of Ca2 (closed state)
Sample
  • Complex: Dimeric lipid scramblase nhTMEM16
    • Protein or peptide: Lipid scramblase nhTMEM16
Keywordsmembrane protein / lipid scramblase / TMEM16 / LIPID TRANSPORT
Function / homology
Function and homology information


cortical endoplasmic reticulum / chloride channel activity / metal ion binding / identical protein binding / membrane
Similarity search - Function
: / Alpha-beta plait domain in TMEM16 lipid scramblase / Anoctamin / : / Calcium-activated chloride channel
Similarity search - Domain/homology
Plasma membrane channel protein
Similarity search - Component
Biological speciesFusarium vanettenii 77-13-4 (fungus)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.64 Å
AuthorsFeng Z / Accardi A
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM106717 United States
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Structural basis of closed groove scrambling by a TMEM16 protein.
Authors: Zhang Feng / Omar E Alvarenga / Alessio Accardi /
Abstract: Activation of Ca-dependent TMEM16 scramblases induces phosphatidylserine externalization, a key step in multiple signaling processes. Current models suggest that the TMEM16s scramble lipids by ...Activation of Ca-dependent TMEM16 scramblases induces phosphatidylserine externalization, a key step in multiple signaling processes. Current models suggest that the TMEM16s scramble lipids by deforming the membrane near a hydrophilic groove and that Ca dependence arises from the different association of lipids with an open or closed groove. However, the molecular rearrangements underlying groove opening and how lipids reorganize outside the closed groove remain unknown. Here we directly visualize how lipids associate at the closed groove of Ca-bound fungal nhTMEM16 in nanodiscs using cryo-EM. Functional experiments pinpoint lipid-protein interaction sites critical for closed groove scrambling. Structural and functional analyses suggest groove opening entails the sequential appearance of two π-helical turns in the groove-lining TM6 helix and identify critical rearrangements. Finally, we show that the choice of scaffold protein and lipids affects the conformations of nhTMEM16 and their distribution, highlighting a key role of these factors in cryo-EM structure determination.
History
DepositionAug 3, 2023-
Header (metadata) releaseMay 8, 2024-
Map releaseMay 8, 2024-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_41458.png

Downloads & links

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Map

FileDownload / File: emd_41458.map.gz / Format: CCP4 / Size: 13.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationLocal refined monomer map used to construct the composite map of nhTMEM16 in lipid nanodiscs with MSP1E3 scaffold protein in the presence of Ca2 (closed state)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (Col.)
0.83 Å/pix.
x 174 pix.
= 143.55 Å		0.83 Å/pix.
x 126 pix.
= 103.95 Å		0.83 Å/pix.
x 157 pix.
= 129.525 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 0.825 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.1
Minimum - Maximum-11.082679000000001 - 19.747388999999998
Average (Standard dev.)-0.000000000022899 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderZYX
Origin564231
Dimensions126157174
Spacing174126157
CellA: 143.55 Å / B: 103.95 Å / C: 129.525 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: halfmap1

Fileemd_41458_half_map_1.map
Annotationhalfmap1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: halfmap2

Fileemd_41458_half_map_2.map
Annotationhalfmap2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Dimeric lipid scramblase nhTMEM16

EntireName: Dimeric lipid scramblase nhTMEM16
Components
  • Complex: Dimeric lipid scramblase nhTMEM16
    • Protein or peptide: Lipid scramblase nhTMEM16

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Supramolecule #1: Dimeric lipid scramblase nhTMEM16

SupramoleculeName: Dimeric lipid scramblase nhTMEM16 / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Fusarium vanettenii 77-13-4 (fungus)

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Macromolecule #1: Lipid scramblase nhTMEM16

MacromoleculeName: Lipid scramblase nhTMEM16 / type: protein_or_peptide / ID: 1 / Enantiomer: LEVO
Source (natural)Organism: Fusarium vanettenii 77-13-4 (fungus)
SequenceString: GPSNLKDFSQ PGSGQESNFG VDFVIHYKVP AAERDEAEAG FVQLIRALTT VGLATEVRHG ENESLLVFVK VASPDLFAKQ VYRARLGDWL HGVRVSAPHN DIAQALQDEP VVEAERLRLI YLMITKPHNE GGAGVTPTNA KWKHVESIFP LHSHSFNKEW IKKWSSKYTL ...String:
GPSNLKDFSQ PGSGQESNFG VDFVIHYKVP AAERDEAEAG FVQLIRALTT VGLATEVRHG ENESLLVFVK VASPDLFAKQ VYRARLGDWL HGVRVSAPHN DIAQALQDEP VVEAERLRLI YLMITKPHNE GGAGVTPTNA KWKHVESIFP LHSHSFNKEW IKKWSSKYTL EQTDIDNIRD KFGESVAFYF AFLRSYFRFL VIPSAFGFGA WLLLGQFSYL YALLCGLWSV VFFEYWKKQE VDLAVQWGVR GVSSIQQSRP EFEWEHEAED PITGEPVKVY PPMKRVKTQL LQIPFALACV VALGALIVTC NSLEVFINEV YSGPGKQYLG FLPTIFLVIG TPTISGVLMG AAEKLNAMEN YATVDAHDAA LIQKQFVLNF MTSYMALFFT AFVYIPFGHI LHPFLNFWRA TAQTLTFSEK ELPTREFQIN PARISNQMFY FTVTAQIVNF ATEVVVPYIK QQAFQKAKQL KSGSKVQEDH EEEAEFLQRV REECTLEEYD VSGDYREMVM QFGYVAMFSV AWPLAACCFL VNNWVELRSD ALKIAISSRR PIPWRTDSIG PWLTALSFLS WLGSITSSAI VYLCSNSKNG TQGEASPLKA WGLLLSILFA EHFYLVVQLA VRFVLSKLDS PGLQKERKER FQTKKRLLQE NLGQDAAEEA AAPGIEHSEK ITREALEEEA RQASIRGHGT PEEMFWQRQR GMQETIEIGR RMIEQQLAAG KNGKKSAPAV PSEKASA

UniProtKB: Plasma membrane channel protein

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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

Concentration5 mg/mL
BufferpH: 8
Component:
ConcentrationNameFormula
50.0 mMTris
150.0 mMKCl
0.5 mMCaCl2
GridMaterial: GOLD / Support film - Material: GOLD
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 288 K / Instrument: FEI VITROBOT MARK IV
DetailsThis sample was monodisperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number real images: 13735 / Average exposure time: 2.0 sec. / Average electron dose: 59.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER / 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

Particle selectionNumber selected: 4574959
Startup modelType of model: NONE
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 2.64 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1.2) / Number images used: 875283
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 3.1.2)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: RELION (ver. 3.1.2)
Final 3D classificationSoftware - Name: RELION (ver. 3.1.2)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6qm9


Chain - Source name: PDB / Chain - Initial model type: experimental model

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