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- EMDB-34998: Cryo-EM structure of human TMEM87A, PE-bound -

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Basic information

Entry
Database: EMDB / ID: EMD-34998
TitleCryo-EM structure of human TMEM87A, PE-bound
Map data
Sample
  • Organelle or cellular component: Transmembrane protein 87A with GFP tag and Twin-strep tag
    • Protein or peptide: Transmembrane protein 87A
  • Ligand: (1S)-2-{[(S)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-1-[(octadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
  • Ligand: CHOLESTEROL
Keywordsnon-selective cation channel / ion channel / membrane protein / Golgi-localized protein
Function / homology
Function and homology information


retrograde transport, endosome to Golgi / Golgi cisterna membrane / RHOA GTPase cycle / ruffle / cellular response to mechanical stimulus / Golgi membrane / Golgi apparatus / plasma membrane / cytosol
Similarity search - Function
Transmembrane protein GPR107/GPR108-like / GOST, seven transmembrane domain
Similarity search - Domain/homology
Transmembrane protein 87A
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsHan A / Kim HM
Funding support Korea, Republic Of, 1 items
OrganizationGrant numberCountry
Other governmentIBS-R030-C1 Korea, Republic Of
CitationJournal: To Be Published
Title: Cryo-EM structure of human TMEM87A, PE-bound
Authors: Han A / Kim HM
History
DepositionDec 19, 2022-
Header (metadata) releaseDec 27, 2023-
Map releaseDec 27, 2023-
UpdateDec 27, 2023-
Current statusDec 27, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_34998.png

Downloads & links

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Map

FileDownload / File: emd_34998.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 0.849 Å
Density
Contour LevelBy AUTHOR: 0.101
Minimum - Maximum-0.32946664 - 0.7027863
Average (Standard dev.)-0.0005385601 (±0.011062544)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 271.68 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_34998_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_34998_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Transmembrane protein 87A with GFP tag and Twin-strep tag

EntireName: Transmembrane protein 87A with GFP tag and Twin-strep tag
Components
  • Organelle or cellular component: Transmembrane protein 87A with GFP tag and Twin-strep tag
    • Protein or peptide: Transmembrane protein 87A
  • Ligand: (1S)-2-{[(S)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-1-[(octadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
  • Ligand: CHOLESTEROL

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Supramolecule #1: Transmembrane protein 87A with GFP tag and Twin-strep tag

SupramoleculeName: Transmembrane protein 87A with GFP tag and Twin-strep tag
type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: #1
Details: Transmembrane protein 87A with GFP tag and Twin-strep tag purified with detergent LMNG/CHS
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Transmembrane protein 87A

MacromoleculeName: Transmembrane protein 87A / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 63.494766 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MAAAAWLQVL PVILLLLGAH PSPLSFFSAG PATVAAADRS KWHIPIPSGK NYFSFGKILF RNTTIFLKFD GEPCDLSLNI TWYLKSADC YNEIYNFKAE EVELYLEKLK EKRGLSGKYQ TSSKLFQNCS ELFKTQTFSG DFMHRLPLLG EKQEAKENGT N LTFIGDKT ...String:
MAAAAWLQVL PVILLLLGAH PSPLSFFSAG PATVAAADRS KWHIPIPSGK NYFSFGKILF RNTTIFLKFD GEPCDLSLNI TWYLKSADC YNEIYNFKAE EVELYLEKLK EKRGLSGKYQ TSSKLFQNCS ELFKTQTFSG DFMHRLPLLG EKQEAKENGT N LTFIGDKT AMHEPLQTWQ DAPYIFIVHI GISSSKESSK ENSLSNLFTM TVEVKGPYEY LTLEDYPLMI FFMVMCIVYV LF GVLWLAW SACYWRDLLR IQFWIGAVIF LGMLEKAVFY AEFQNIRYKG ESVQGALILA ELLSAVKRSL ARTLVIIVSL GYG IVKPRL GVTLHKVVVA GALYLLFSGM EGVLRVTGAQ TDLASLAFIP LAFLDTALCW WIFISLTQTM KLLKLRRNIV KLSL YRHFT NTLILAVAAS IVFIIWTTMK FRIVTCQSDW RELWVDDAIW RLLFSMILFV IMVLWRPSAN NQRFAFSPLS EEEEE DEQK EPMLKESFEG MKMRSTKQEP NGNSKVNKAQ EDDLKWVEEN VPSSVTDVAL PALLDSDEER MITHFERSKM E

UniProtKB: Transmembrane protein 87A

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Macromolecule #4: (1S)-2-{[(S)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-1-[(octadeca...

MacromoleculeName: (1S)-2-{[(S)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-1-[(octadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate
type: ligand / ID: 4 / Number of copies: 1 / Formula: L9Q
Molecular weightTheoretical: 746.05 Da
Chemical component information

ChemComp-L9Q:
(1S)-2-{[(S)-(2-aminoethoxy)(hydroxy)phosphoryl]oxy}-1-[(octadecanoyloxy)methyl]ethyl (9Z)-octadec-9-enoate

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Macromolecule #5: CHOLESTEROL

MacromoleculeName: CHOLESTEROL / type: ligand / ID: 5 / Number of copies: 2 / Formula: CLR
Molecular weightTheoretical: 386.654 Da
Chemical component information

ChemComp-CLR:
CHOLESTEROL

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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

Concentration0.8 mg/mL
BufferpH: 9
Component:
ConcentrationFormulaName
50.0 mMTristrisaminomethane
250.0 mMNaClSodium Chloride
0.01 % (w/v)DDMDodecylmaltoside
0.002 % (w/v)CHSCholesteryl hemisuccinate

Details: 50mM Tris pH 9.0, 250mM NaCl, 0.01% (w/v) LMNG, 0.002% (w/v) CHS
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 75 sec. / Details: 15mA
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
DetailsThis sample was monodisperse

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Alignment procedureComa free - Residual tilt: 10.0 mrad
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
DetailsUsing Zemlin tableau in sherpa
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 10377 / Average exposure time: 6.14 sec. / Average electron dose: 67.72 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated magnification: 58900 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.9000000000000001 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 105000
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: 8101104
Details: 96,330 particles were first picked using a blob picker of cryoSPARC. 2D class average images were generated as templates for subsequent reference-based auto-picking.
Startup modelType of model: OTHER / Details: AlphaFoldDB: Q8NBN3
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.3.2) / Software - details: Local refinement
Details: Non-uniform refinement and CTF refinement were used to improve the particle alignment and map quality.
Number images used: 445198
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: cryoSPARC (ver. 3.3.2) / Software - details: Ab-intio reconstruction
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2) / Software - details: Non-uniform refinement
Final 3D classificationNumber classes: 6 / Avg.num./class: 451461 / Software - Name: cryoSPARC (ver. 3.3.2) / Software - details: Hetero refinement
FSC plot (resolution estimation)

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Atomic model buiding 1

DetailsChimera Fit in map tool was used for initial local fitting. Then, Real-space refinement with the rigid body option in PHENIX was used for flexible fitting.
RefinementSpace: REAL / Protocol: RIGID BODY FIT / Overall B value: 40
Output model

PDB-8hsi:
Cryo-EM structure of human TMEM87A, PE-bound

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