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- EMDB-38731: Cryo-EM structure of the aspartate:alanine antiporter AspT mutant L60C -

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Basic information

Entry
Database: EMDB / ID: EMD-38731
TitleCryo-EM structure of the aspartate:alanine antiporter AspT mutant L60C
Map data
Sample
  • Organelle or cellular component: aspartate:alanine antiporter AspT mutant - L60C
    • Protein or peptide: Aspartate/alanine antiporter
Keywordsamino acid / antiporter / elevator mechanism / TRANSPORT PROTEIN
Function / homologyYidE/YbjL duplication / Aspartate-alanine antiporter / Predicted Permease Membrane Region / : / antiporter activity / identical protein binding / plasma membrane / Aspartate/alanine antiporter
Function and homology information
Biological speciesTetragenococcus halophilus (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.56 Å
AuthorsNanatani K / Kanno R / Kawabata T / Watanabe S / Hidaka M / Yamanaka T / Toda K / Fujiki T / Kunii K / Miyamoto A ...Nanatani K / Kanno R / Kawabata T / Watanabe S / Hidaka M / Yamanaka T / Toda K / Fujiki T / Kunii K / Miyamoto A / Chiba F / Ogasawara S / Murata T / Humbel BM / Inaba K / Mitsuoka K / Guan L / Abe K / Yamamoto M / Koshiba S
Funding support Japan, 5 items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)16K07653 Japan
Japan Society for the Promotion of Science (JSPS)20K05779 Japan
Japan Society for the Promotion of Science (JSPS)23K05004 Japan
Japan Agency for Medical Research and Development (AMED)JP23ama121019 Japan
Japan Agency for Medical Research and Development (AMED)JP23ama121038 Japan
CitationJournal: To Be Published
Title: Cryo-EM structure and molecular mechanism of the aspartate:alanine antiporter AspT from Tetragenococcus halophilus
Authors: Nanatani K / Kanno R / Kawabata T / Watanabe S / Hidaka M / Yamanaka T / Toda K / Fujiki T / Kunii K / Miyamoto A / Chiba F / Ogasawara S / Murata T / Humbel BM / Inaba K / Mitsuoka K / Guan ...Authors: Nanatani K / Kanno R / Kawabata T / Watanabe S / Hidaka M / Yamanaka T / Toda K / Fujiki T / Kunii K / Miyamoto A / Chiba F / Ogasawara S / Murata T / Humbel BM / Inaba K / Mitsuoka K / Guan L / Abe K / Yamamoto M / Koshiba S
History
DepositionJan 16, 2024-
Header (metadata) releaseJan 22, 2025-
Map releaseJan 22, 2025-
UpdateJan 22, 2025-
Current statusJan 22, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_38731.png

Downloads & links

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Map

FileDownload / File: emd_38731.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.79 Å/pix.
x 256 pix.
= 201.728 Å		0.79 Å/pix.
x 256 pix.
= 201.728 Å		0.79 Å/pix.
x 256 pix.
= 201.728 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.788 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.193
Minimum - Maximum-1.0394294 - 1.3563445
Average (Standard dev.)0.0017518044 (±0.035531424)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 201.728 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_38731_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_38731_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 : aspartate:alanine antiporter AspT mutant - L60C

EntireName: aspartate:alanine antiporter AspT mutant - L60C
Components
  • Organelle or cellular component: aspartate:alanine antiporter AspT mutant - L60C
    • Protein or peptide: Aspartate/alanine antiporter

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Supramolecule #1: aspartate:alanine antiporter AspT mutant - L60C

SupramoleculeName: aspartate:alanine antiporter AspT mutant - L60C / type: organelle_or_cellular_component / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Tetragenococcus halophilus (bacteria) / Strain: D10
Molecular weightTheoretical: 57.9 KDa

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Macromolecule #1: Aspartate/alanine antiporter

MacromoleculeName: Aspartate/alanine antiporter / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Tetragenococcus halophilus (bacteria) / Strain: D10
Molecular weightTheoretical: 57.148918 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MNAIGNFLVG TPVFTIFIAL ALGYLLGKLK IGSFTLGATV GVLIVALLIG QLGVFPRDTC LGDIFFDFFM FAIGYRVGPS FISSMKKFG AKIVYATLIF LVSAFIVAYA AFKMFHIGPG IAAGIIAGGL TQSAVIGSSL ETISKLPISD HLKTLYSNQI P IVYTLTYV ...String:
MNAIGNFLVG TPVFTIFIAL ALGYLLGKLK IGSFTLGATV GVLIVALLIG QLGVFPRDTC LGDIFFDFFM FAIGYRVGPS FISSMKKFG AKIVYATLIF LVSAFIVAYA AFKMFHIGPG IAAGIIAGGL TQSAVIGSSL ETISKLPISD HLKTLYSNQI P IVYTLTYV FGTIGVLIFL RDIMPKLMHI DLKKQAVKTA KELDMIPVPV IVASTHFYTI NDGSSLIGQT LGTVNTKFAK GL VAAGLND SADMASVINA GDVLAISGGI DEIGRAVQEF NLLEVTGKTK AYVSKQVVLK KNFSADVLKN AQDKGVLVAT LAG DVMDPA QFSTLKPAES VTLVGQKDAV SEVQSQLGRL RAAENIINYS WFALGIALSA ALGIVGTKVS GVPIALGGGT ASLI VGLVQ SIYRDKHAHM DTIPDSLLEF FQSIGLNLFI ATVGLSAAKT FISAIQSMGI SVLLIGAVIS ILPHIITFVI AYYLM KMEP ISIIGAQTGA DTLSAALNDV SERVGSDASP FFAAAVAPAY AIGNIFLTLM GPIFIVLLS

UniProtKB: Aspartate/alanine antiporter

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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

Concentration6.14 mg/mL
BufferpH: 6
Component:
ConcentrationFormulaName
20.0 mMMES2-(N-morpholino)ethanesulfonic acid
100.0 mMNaClsodium chloride
0.003 %LMNGLauryl maltose neopentyl glycol
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeJEOL CRYO ARM 300
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 6000 / 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: 2.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 60000
Sample stageSpecimen holder model: JEOL CRYOSPECPORTER / Cooling holder cryogen: NITROGEN

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Image processing

Particle selectionNumber selected: 1634276
Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C2 (2 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.56 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 250605
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC
Final 3D classificationNumber classes: 2 / Software - Name: cryoSPARC

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
RefinementSpace: REAL / Protocol: BACKBONE TRACE
Output model

PDB-8xw5:
Cryo-EM structure of the aspartate:alanine antiporter AspT mutant L60C

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