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- EMDB-60461: AtALMT9 with LMNG and sterol mimic CHS (sterol2 class) -

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Basic information

Entry
Database: EMDB / ID: EMD-60461
TitleAtALMT9 with LMNG and sterol mimic CHS (sterol2 class)
Map data
Sample
  • Complex: AtALMT9 with LMNG and sterol mimic CHS
    • Protein or peptide: Aluminum-activated malate transporter 9
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: D-MALATE
  • Ligand: 1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE
  • Ligand: water
KeywordsPlant / Stomata / Vacuole / ALMT / Ion channel / MEMBRANE PROTEIN
Function / homologymalate transport / Aluminum-activated malate transporter / Aluminium activated malate transporter / plant-type vacuole membrane / monoatomic anion channel activity / Aluminum-activated malate transporter 9
Function and homology information
Biological speciesArabidopsis thaliana (thale cress)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.95 Å
AuthorsLee Y / Lee S
Funding support Korea, Republic Of, 3 items
OrganizationGrant numberCountry
National Research Foundation (NRF, Korea)2022R1A2C100988211 Korea, Republic Of
National Research Foundation (NRF, Korea)RS-2023-00223552 Korea, Republic Of
National Research Foundation (NRF, Korea)2021M3A9I4022936 Korea, Republic Of
CitationJournal: Nat Commun / Year: 2025
Title: Structural basis for malate-driven, pore lipid-regulated activation of the Arabidopsis vacuolar anion channel ALMT9.
Authors: Yeongmok Lee / Elsa Demes-Causse / Jaemin Yoo / Seo Young Jang / Seoyeon Jung / Justyna Jaślan / Geum-Sook Hwang / Jejoong Yoo / Alexis De Angeli / Sangho Lee /
Abstract: In plant cells, ALMTs are key plasma and vacuolar membrane-localized anion channels regulating plant responses to the environment. Vacuolar ALMTs control anion accumulation in plant cells and, in ...In plant cells, ALMTs are key plasma and vacuolar membrane-localized anion channels regulating plant responses to the environment. Vacuolar ALMTs control anion accumulation in plant cells and, in guard cells, they regulate stomata aperture. The activation of vacuolar ALMTs depends on voltage and cytosolic malate, but the underlying molecular mechanisms remain elusive. Here we report the cryo-EM structures of ALMT9 from Arabidopsis thaliana (AtALMT9), a malate-activated vacuolar anion channel, in plugged and unplugged lipid-bound states. In all these states, membrane lipids interact with the ion conduction pathway of AtALMT9. We identify two unplugged states presenting two distinct pore width profiles. Combining structural and functional analysis we identified conserved residues involved in ion conduction and in the pore lipid interaction. Molecular dynamics simulations revealed a peculiar anion conduction mechanism in AtALMT9. We propose a voltage-dependent activation mechanism based on the competition between pore lipids and malate at the cytosolic entrance of the channel.
History
DepositionJun 7, 2024-
Header (metadata) releaseMar 12, 2025-
Map releaseMar 12, 2025-
UpdateMar 12, 2025-
Current statusMar 12, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_60461.png

Downloads & links

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Map

FileDownload / File: emd_60461.map.gz / Format: CCP4 / Size: 38.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
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AxesZ (Sec.)Y (Row.)X (Col.)
0.85 Å/pix.
x 216 pix.
= 183.6 Å		0.85 Å/pix.
x 216 pix.
= 183.6 Å		0.85 Å/pix.
x 216 pix.
= 183.6 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.85 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.29
Minimum - Maximum-1.4967973 - 2.239523
Average (Standard dev.)0.0020752128 (±0.06906992)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions216216216
Spacing216216216
CellA=B=C: 183.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_60461_half_map_1.map
Projections & Slices
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Histogram (log scale)

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Half map: #1

Fileemd_60461_half_map_2.map
Projections & Slices
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Sample components

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Entire : AtALMT9 with LMNG and sterol mimic CHS

EntireName: AtALMT9 with LMNG and sterol mimic CHS
Components
  • Complex: AtALMT9 with LMNG and sterol mimic CHS
    • Protein or peptide: Aluminum-activated malate transporter 9
  • Ligand: CHOLESTEROL HEMISUCCINATE
  • Ligand: D-MALATE
  • Ligand: 1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE
  • Ligand: water

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Supramolecule #1: AtALMT9 with LMNG and sterol mimic CHS

SupramoleculeName: AtALMT9 with LMNG and sterol mimic CHS / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 130 KDa

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Macromolecule #1: Aluminum-activated malate transporter 9

MacromoleculeName: Aluminum-activated malate transporter 9 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 67.121008 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: MAAKQGSFRH GILEKRERLL SNNGFSDFRF TDIESNDLLE NENCGRRTRL CCCCSCGNLS EKISGVYDDA KDVARKAWEM GVSDPRKIV FSAKIGLALT IVALLIFYQE PNPDLSRYSV WAILTVVVVF EFTIGATLSK GFNRALGTLS AGGLALGMAE L STLFGDWE ...String:
MAAKQGSFRH GILEKRERLL SNNGFSDFRF TDIESNDLLE NENCGRRTRL CCCCSCGNLS EKISGVYDDA KDVARKAWEM GVSDPRKIV FSAKIGLALT IVALLIFYQE PNPDLSRYSV WAILTVVVVF EFTIGATLSK GFNRALGTLS AGGLALGMAE L STLFGDWE EIFCTLSIFC IGFLATFMKL YPSMKAYEYG FRVFLLTYCY ILISGFRTGQ FIEVAISRFL LIALGAGVSL GV NMFIYPI WAGEDLHNLV VKNFMNVATS LEGCVNGYLR CLEYERIPSK ILTYQASEDP VYKGYRSAVE STSQEESLMS FAI WEPPHG PYKSFNYPWK NYVKLSGALK HCAFTVMALH GCILSEIQAP EERRQVFRQE LQRVGVEGAK LLRELGEKVK KMEK LGPVD LLFEVHLAAE ELQHKIDKKS YLLVNSECWE IGNRATKESE PQELLSLEDS DPPENHAPPI YAFKSLSEAV LEIPP SWGE KNHREALNHR PTFSKQVSWP ARLVLPPHLE TTNGASPLVE TTKTYESASA LSLATFASLL IEFVARLQNV VDAFKE LSQ KANFKEPEIV TTGTDVEFSG ERVGLGQKIR RCFGM

UniProtKB: Aluminum-activated malate transporter 9

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Macromolecule #2: CHOLESTEROL HEMISUCCINATE

MacromoleculeName: CHOLESTEROL HEMISUCCINATE / type: ligand / ID: 2 / Number of copies: 6 / Formula: Y01
Molecular weightTheoretical: 486.726 Da
Chemical component information

ChemComp-Y01:
CHOLESTEROL HEMISUCCINATE

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Macromolecule #3: D-MALATE

MacromoleculeName: D-MALATE / type: ligand / ID: 3 / Number of copies: 1 / Formula: MLT
Molecular weightTheoretical: 134.087 Da
Chemical component information

ChemComp-MLT:
D-MALATE

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Macromolecule #4: 1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE

MacromoleculeName: 1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE / type: ligand / ID: 4 / Number of copies: 1 / Formula: 3PH
Molecular weightTheoretical: 704.998 Da
Chemical component information

ChemComp-3PH:
1,2-DIACYL-GLYCEROL-3-SN-PHOSPHATE

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Macromolecule #5: water

MacromoleculeName: water / type: ligand / ID: 5 / Number of copies: 8 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
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

Concentration1.9 mg/mL
BufferpH: 7.6
Component:
ConcentrationFormulaName
20.0 mMC8H18N2O4SHEPES
150.0 mMNaClSodium chloride
1.0 mMC9H15O6PTCEP
0.005 % (w/v)C47H88O22LMNG
0.001 % (w/v)C31H50O4CHS
10.0 mMC4H6O5Malate
GridModel: Au-flat 1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY / Support film - Film thickness: 45 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 2 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 14 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.9000000000000001 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1648820
Startup modelType of model: NONE
Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C2 (2 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 2.95 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.0) / Number images used: 190859
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0) / Details: NU-refinement
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.0) / Details: NU-refinement
Final 3D classificationNumber classes: 4 / Avg.num./class: 47847 / Software - Name: cryoSPARC (ver. 4.0)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
Output model

PDB-8ztg:
AtALMT9 with LMNG and sterol mimic CHS (sterol2 class)

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