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- EMDB-18653: Cryo-EM structure of the FB-bound yeast Ceramide Synthase -

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Basic information

Entry
Database: EMDB / ID: EMD-18653
TitleCryo-EM structure of the FB-bound yeast Ceramide Synthase
Map data
Sample
  • Complex: Heterotetrameric complex of yeast ceramide synthase with Lag1, Lac1 and Lip1
    • Protein or peptide: Ceramide synthase LAG1
    • Protein or peptide: Ceramide synthase LAC1
    • Protein or peptide: Ceramide synthase subunit LIP1
  • Ligand: 1,2-Distearoyl-sn-glycerophosphoethanolamine
  • Ligand: Macrofusine
  • Ligand: hexacosanoic acid
  • Ligand: [(2S)-1-hexadecanoyloxy-3-[hydroxy-[(2S,3R,5S,6R)-2,3,4,5,6-pentahydroxycyclohexyl]oxy-phosphoryl]oxy-propan-2-yl] heptadecanoate
  • Ligand: AMMONIUM ION
KeywordsCeramide Synthase / MEMBRANE PROTEIN
Function / homology
Function and homology information


very-long-chain ceramide synthase / acyl-CoA ceramide synthase complex / Sphingolipid de novo biosynthesis / sphingosine N-acyltransferase activity / ceramide biosynthetic process / nuclear periphery / nuclear envelope / endoplasmic reticulum membrane / endoplasmic reticulum
Similarity search - Function
TRAM/LAG1/CLN8 homology domain / Sphingosine N-acyltransferase Lag1/Lac1-like / TLC domain / TLC domain profile. / TRAM, LAG1 and CLN8 homology domains.
Similarity search - Domain/homology
Ceramide synthase LAC1 / Ceramide synthase LAG1 / Ceramide synthase subunit LIP1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsSchaefer J / Clausmeyer L / Koerner C / Moeller A / Froehlich F
Funding support Germany, 1 items
OrganizationGrant numberCountry
German Research Foundation (DFG)SFB1557 Germany
CitationJournal: Nat Struct Mol Biol / Year: 2025
Title: Structure of the yeast ceramide synthase.
Authors: Jan-Hannes Schäfer / Lena Clausmeyer / Carolin Körner / Bianca M Esch / Verena N Wolf / Jennifer Sapia / Yara Ahmed / Stefan Walter / Stefano Vanni / Dovile Januliene / Arne Moeller / Florian Fröhlich /
Abstract: Ceramides are essential lipids involved in forming complex sphingolipids and acting as signaling molecules. They result from the N-acylation of a sphingoid base and a CoA-activated fatty acid, a ...Ceramides are essential lipids involved in forming complex sphingolipids and acting as signaling molecules. They result from the N-acylation of a sphingoid base and a CoA-activated fatty acid, a reaction catalyzed by the ceramide synthase (CerS) family of enzymes. Yet, the precise structural details and catalytic mechanisms of CerSs have remained elusive. Here we used cryo-electron microscopy single-particle analysis to unravel the structure of the yeast CerS complex in both an active and a fumonisin B1-inhibited state. Our results reveal the complex's architecture as a dimer of Lip1 subunits bound to the catalytic subunits Lag1 and Lac1. Each catalytic subunit forms a hydrophobic crevice connecting the cytosolic site with the intermembrane space. The active site, located centrally in the tunnel, was resolved in a substrate preloaded state, representing one intermediate in ceramide synthesis. Our data provide evidence for competitive binding of fumonisin B1 to the acyl-CoA-binding tunnel.
History
DepositionOct 13, 2023-
Header (metadata) releaseOct 23, 2024-
Map releaseOct 23, 2024-
UpdateJul 9, 2025-
Current statusJul 9, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_18653.png

Downloads & links

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Map

FileDownload / File: emd_18653.map.gz / Format: CCP4 / Size: 347.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.68 Å/pix.
x 450 pix.
= 306. Å		0.68 Å/pix.
x 450 pix.
= 306. Å		0.68 Å/pix.
x 450 pix.
= 306. Å

Surface

Projections

Slices (1/3)

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Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.68 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.12
Minimum - Maximum-0.61564714 - 1.0430472
Average (Standard dev.)0.00026234167 (±0.0175536)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions450450450
Spacing450450450
CellA=B=C: 306.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_18653_msk_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_18653_half_map_1.map
Projections & Slices
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Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_18653_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Heterotetrameric complex of yeast ceramide synthase with Lag1, La...

EntireName: Heterotetrameric complex of yeast ceramide synthase with Lag1, Lac1 and Lip1
Components
  • Complex: Heterotetrameric complex of yeast ceramide synthase with Lag1, Lac1 and Lip1
    • Protein or peptide: Ceramide synthase LAG1
    • Protein or peptide: Ceramide synthase LAC1
    • Protein or peptide: Ceramide synthase subunit LIP1
  • Ligand: 1,2-Distearoyl-sn-glycerophosphoethanolamine
  • Ligand: Macrofusine
  • Ligand: hexacosanoic acid
  • Ligand: [(2S)-1-hexadecanoyloxy-3-[hydroxy-[(2S,3R,5S,6R)-2,3,4,5,6-pentahydroxycyclohexyl]oxy-phosphoryl]oxy-propan-2-yl] heptadecanoate
  • Ligand: AMMONIUM ION

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Supramolecule #1: Heterotetrameric complex of yeast ceramide synthase with Lag1, La...

SupramoleculeName: Heterotetrameric complex of yeast ceramide synthase with Lag1, Lac1 and Lip1
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 140 KDa

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Macromolecule #1: Ceramide synthase LAG1

MacromoleculeName: Ceramide synthase LAG1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 37.150789 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: YREMNYRHSW LTPFFILVCV YSAYFLSGNR TESNPLHMFV AISYQVDGTD SYAKGIKDLS FVFFYMIFFT FLREFLMDVV IRPFTVYLN VTSEHRQKRM LEQMYAIFYC GVSGPFGLYI MYHSDLWLFK TKPMYRTYPV ITNPFLFKIF YLGQAAFWAQ Q ACVLVLQL ...String:
YREMNYRHSW LTPFFILVCV YSAYFLSGNR TESNPLHMFV AISYQVDGTD SYAKGIKDLS FVFFYMIFFT FLREFLMDVV IRPFTVYLN VTSEHRQKRM LEQMYAIFYC GVSGPFGLYI MYHSDLWLFK TKPMYRTYPV ITNPFLFKIF YLGQAAFWAQ Q ACVLVLQL EKPRKDYKEL VFHHIVTLLL IWSSYVFHFT KMGLAIYITM DVSDFFLSLS KTLNYLNSVF TPFVFGLFVF FW IYLRHVV NIRILWSVLT EFRHEGNYVL NFATQQYKCW ISLPIVFVLI AALQLVNLYW LFLILRILYR LI

UniProtKB: Ceramide synthase LAG1

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Macromolecule #2: Ceramide synthase LAC1

MacromoleculeName: Ceramide synthase LAC1 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 35.802074 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString: ISYRHAWIAP LMILIAVYSA YFTSGNTTKT NVLHRFVAVS YQIGDTNAYG KGINDLCFVF YYMIFFTFLR EFLMDVVIRP FAIRLHVTS KHRIKRIMEQ MYAIFYTGVS GPFGIYCMYH SDLWFFNTKA MYRTYPDFTN PFLFKVFYLG QAAFWAQQAC I LVLQLEKP ...String:
ISYRHAWIAP LMILIAVYSA YFTSGNTTKT NVLHRFVAVS YQIGDTNAYG KGINDLCFVF YYMIFFTFLR EFLMDVVIRP FAIRLHVTS KHRIKRIMEQ MYAIFYTGVS GPFGIYCMYH SDLWFFNTKA MYRTYPDFTN PFLFKVFYLG QAAFWAQQAC I LVLQLEKP RKDHNELTFH HIVTLLLIWS SYVFHFTKMG LPIYITMDVS DFLLSFSKTL NYLDSGLAFF SFAIFVVAWI YL RHYINLK ILWSVLTQFR TEGNYVLNFA TQQYKCWISL PIVFVLIGAL QLVNLYWLFL IFRVL

UniProtKB: Ceramide synthase LAC1

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Macromolecule #3: Ceramide synthase subunit LIP1

MacromoleculeName: Ceramide synthase subunit LIP1 / type: protein_or_peptide / ID: 3 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 15.347415 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)
SequenceString:
KIFNLFRVCF ISLLLIAAVE YFKYGTRINY EWFHCTPIKE PQSGSVIKLW ARGGPSCDKR GEYKTIVKRI TRDYEPNDEH LSFCIIEND NVPPVHYPIH EDKGEPGYVA YVGYDTDSEL VQELCADSTI YHM

UniProtKB: Ceramide synthase subunit LIP1

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Macromolecule #4: 1,2-Distearoyl-sn-glycerophosphoethanolamine

MacromoleculeName: 1,2-Distearoyl-sn-glycerophosphoethanolamine / type: ligand / ID: 4 / Number of copies: 4 / Formula: 3PE
Molecular weightTheoretical: 748.065 Da
Chemical component information

ChemComp-3PE:
1,2-Distearoyl-sn-glycerophosphoethanolamine / phospholipid*YM

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Macromolecule #5: Macrofusine

MacromoleculeName: Macrofusine / type: ligand / ID: 5 / Number of copies: 2 / Formula: WXE
Molecular weightTheoretical: 721.83 Da

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Macromolecule #6: hexacosanoic acid

MacromoleculeName: hexacosanoic acid / type: ligand / ID: 6 / Number of copies: 2 / Formula: 7PO
Molecular weightTheoretical: 396.69 Da
Chemical component information

ChemComp-7PO:
hexacosanoic acid

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Macromolecule #7: [(2S)-1-hexadecanoyloxy-3-[hydroxy-[(2S,3R,5S,6R)-2,3,4,5,6-penta...

MacromoleculeName: [(2S)-1-hexadecanoyloxy-3-[hydroxy-[(2S,3R,5S,6R)-2,3,4,5,6-pentahydroxycyclohexyl]oxy-phosphoryl]oxy-propan-2-yl] heptadecanoate
type: ligand / ID: 7 / Number of copies: 2 / Formula: PIJ
Molecular weightTheoretical: 825.059 Da

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Macromolecule #8: AMMONIUM ION

MacromoleculeName: AMMONIUM ION / type: ligand / ID: 8 / Number of copies: 2 / Formula: NH4
Molecular weightTheoretical: 18.038 Da
Chemical component information

ChemComp-NH4:
AMMONIUM ION

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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: 6.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS GLACIOS
Specialist opticsEnergy filter - Name: TFS Selectris
Image recordingFilm or detector model: TFS FALCON 4i (4k x 4k) / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.8 µm

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: INSILICO MODEL
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 259645
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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