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- EMDB-24731: afTMEM16 in C18 lipid nanodiscs with MSP1E3 scaffold protein in t... -

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Basic information

Entry
Database: EMDB / ID: EMD-24731
TitleafTMEM16 in C18 lipid nanodiscs with MSP1E3 scaffold protein in the presence of Ca2+, monomer with extra lipids
Map dataunsharpened map from a focused reconstruction on one monomer from symmetry expanded particles, used to build additional lipids.
Sample
  • Complex: afTMEM16 scramblase C18 lipid nanodiscs with MSP1E3 scaffold in the presence of Ca2+, monomer with additional lipids
    • Protein or peptide: afTMEM16 lipid scramblase
  • Ligand: CALCIUM IONCalcium
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
  • Ligand: water
Function / homology
Function and homology information


phospholipid scramblase activity / cortical endoplasmic reticulum / phospholipid translocation / chloride channel activity / voltage-gated calcium channel activity / chloride transmembrane transport / monoatomic ion transmembrane transport / 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 (Aqy1), putative
Similarity search - Component
Biological speciesAspergillus fumigatus (mold) / Neosartorya fumigata (strain CEA10 / CBS 144.89 / FGSC A1163) (mold)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.3 Å
AuthorsFalzone ME / 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 Commun / Year: 2022
Title: TMEM16 scramblases thin the membrane to enable lipid scrambling.
Authors: Maria E Falzone / Zhang Feng / Omar E Alvarenga / Yangang Pan / ByoungCheol Lee / Xiaolu Cheng / Eva Fortea / Simon Scheuring / Alessio Accardi /
Abstract: TMEM16 scramblases dissipate the plasma membrane lipid asymmetry to activate multiple eukaryotic cellular pathways. Scrambling was proposed to occur with lipid headgroups moving between leaflets ...TMEM16 scramblases dissipate the plasma membrane lipid asymmetry to activate multiple eukaryotic cellular pathways. Scrambling was proposed to occur with lipid headgroups moving between leaflets through a membrane-spanning hydrophilic groove. Direct information on lipid-groove interactions is lacking. We report the 2.3 Å resolution cryogenic electron microscopy structure of the nanodisc-reconstituted Ca-bound afTMEM16 scramblase showing how rearrangement of individual lipids at the open pathway results in pronounced membrane thinning. Only the groove's intracellular vestibule contacts lipids, and mutagenesis suggests scrambling does not require specific protein-lipid interactions with the extracellular vestibule. We find scrambling can occur outside a closed groove in thinner membranes and is inhibited in thicker membranes, despite an open pathway. Our results show afTMEM16 thins the membrane to enable scrambling and that an open hydrophilic pathway is not a structural requirement to allow rapid transbilayer movement of lipids. This mechanism could be extended to other scramblases lacking a hydrophilic groove.
History
DepositionAug 23, 2021-
Header (metadata) releaseMay 18, 2022-
Map releaseMay 18, 2022-
UpdateMay 25, 2022-
Current statusMay 25, 2022Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_24731.png

Downloads & links

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Map

FileDownload / File: emd_24731.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationunsharpened map from a focused reconstruction on one monomer from symmetry expanded particles, used to build additional lipids.
Voxel sizeX=Y=Z: 0.7067 Å
Density
Contour LevelBy AUTHOR: 0.0049
Minimum - Maximum-0.014968073 - 0.03991337
Average (Standard dev.)-1.6320157e-05 (±0.0009940219)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 271.3728 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_24731_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: unsharpened map from a focused reconstruction on one...

Fileemd_24731_additional_1.map
Annotationunsharpened map from a focused reconstruction on one monomer from symmetry expanded particles
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : afTMEM16 scramblase C18 lipid nanodiscs with MSP1E3 scaffold in t...

EntireName: afTMEM16 scramblase C18 lipid nanodiscs with MSP1E3 scaffold in the presence of Ca2+, monomer with additional lipids
Components
  • Complex: afTMEM16 scramblase C18 lipid nanodiscs with MSP1E3 scaffold in the presence of Ca2+, monomer with additional lipids
    • Protein or peptide: afTMEM16 lipid scramblase
  • Ligand: CALCIUM IONCalcium
  • Ligand: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
  • Ligand: water

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Supramolecule #1: afTMEM16 scramblase C18 lipid nanodiscs with MSP1E3 scaffold in t...

SupramoleculeName: afTMEM16 scramblase C18 lipid nanodiscs with MSP1E3 scaffold in the presence of Ca2+, monomer with additional lipids
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Aspergillus fumigatus (mold)
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)

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Macromolecule #1: afTMEM16 lipid scramblase

MacromoleculeName: afTMEM16 lipid scramblase / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Neosartorya fumigata (strain CEA10 / CBS 144.89 / FGSC A1163) (mold)
Strain: CEA10 / CBS 144.89 / FGSC A1163
Molecular weightTheoretical: 84.616859 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: MAFNPAPKAV QENHHVDYVI RFNYGDIDTP EAIKKFEVLL LELSEVGLQT EVRQGDENSL FVFVRAASKK KLKRAVYQSR VRDWLYGVR NTEPEPASSA KPQSEAERLL VIYHLITVPK AEGGAGITPR HGEWKNVDAI FPLHDEETNR QCMREWSKKT F LSTEDLDR ...String:
MAFNPAPKAV QENHHVDYVI RFNYGDIDTP EAIKKFEVLL LELSEVGLQT EVRQGDENSL FVFVRAASKK KLKRAVYQSR VRDWLYGVR NTEPEPASSA KPQSEAERLL VIYHLITVPK AEGGAGITPR HGEWKNVDAI FPLHDEETNR QCMREWSKKT F LSTEDLDR IRNTFGEHVG FYFAFLQSYF RFLMFPAAFG FSCWLLLGSF SIIYTVVNCL WCIVFIEYWK RQEEDLSCRW QT KGVSAVH EKRAEFKPEK EIRDESTGEV RGVFPATKRM YRQLLQVPFA LLAAVALGAI IATCFAIEIF ISEVYNGPLK GYL VFIPTI LVSALIPTMS AVLLTVATKL NDYENYETQD AYKVALTQKI FVVNFITSYL PIILTAFVYV PFASRIVPYL DVFH LTVRP FVSKEHAIKA RTEFSINPDR LRKQVIYFTV TAQIVGFALE TIVPFVKQRV FREYKEYTKK QHAKAEPGNG AGEKK TVSL GDDEDEARFL TRVRNEAELE DYDVTDDLRE MCIQFGYLAL FSPVWPLVPV SFLINNWVEL RSDFFKICVE CKRPWP QRA DTIGPWLDSL GFLSWVGSIT SSALVYMFSN GHEGPNGEPT TIRCWALLLT IFFSEHLYLI VRYAVRSALA KLEPPNT RR ERIERFMMRK RYLDTVLSAE SDDDADEVKG VVSSIPPSEI TRESLEQDAR DWSKQGTDPT ERFWMRQRGW KESAEVGL S LITKAKGDET KKQQ

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Macromolecule #2: CALCIUM ION

MacromoleculeName: CALCIUM ION / type: ligand / ID: 2 / Number of copies: 2 / Formula: CA
Molecular weightTheoretical: 40.078 Da

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Macromolecule #3: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]o...

MacromoleculeName: (1R)-2-{[(S)-{[(2S)-2,3-dihydroxypropyl]oxy}(hydroxy)phosphoryl]oxy}-1-[(hexadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
type: ligand / ID: 3 / Number of copies: 16 / Formula: PGW
Molecular weightTheoretical: 749.007 Da

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Macromolecule #4: water

MacromoleculeName: water / type: ligand / ID: 4 / Number of copies: 24 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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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
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 42.8227 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.3 Å / Resolution method: FSC 0.143 CUT-OFF
Details: Sorted particles for the C2 dimer reconstruction were symmetry expanded and further classified with a mask on one monomer with and without local alignment with varying T values. These ...Details: Sorted particles for the C2 dimer reconstruction were symmetry expanded and further classified with a mask on one monomer with and without local alignment with varying T values. These 1058829 sorted into a class with strong density for additional lipids.
Number images used: 1058829
FSC plot (resolution estimation)

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