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- EMDB-35862: Cryo-EM structure of the C26-CoA-bound Lac1-Lip1 complex -

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Basic information

Entry
Database: EMDB / ID: EMD-35862
TitleCryo-EM structure of the C26-CoA-bound Lac1-Lip1 complex
Map data
Sample
  • Complex: Lac1-Lip1 complex
    • Protein or peptide: Ceramide synthase LAC1
    • Protein or peptide: Ceramide synthase subunit LIP1
  • Ligand: (4S,7R)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE
  • Ligand: Hexacosanoyl-CoA
KeywordsSubstrate / Complex / Transferase
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 subunit LIP1
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.09 Å
AuthorsXie T / Fang Q / Gong X
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: EMBO J / Year: 2023
Title: Structure and mechanism of a eukaryotic ceramide synthase complex.
Authors: Tian Xie / Qi Fang / Zike Zhang / Yanfei Wang / Feitong Dong / Xin Gong /
Abstract: Ceramide synthases (CerS) catalyze ceramide formation via N-acylation of a sphingoid base with a fatty acyl-CoA and are attractive drug targets for treating numerous metabolic diseases and cancers. ...Ceramide synthases (CerS) catalyze ceramide formation via N-acylation of a sphingoid base with a fatty acyl-CoA and are attractive drug targets for treating numerous metabolic diseases and cancers. Here, we present the cryo-EM structure of a yeast CerS complex, consisting of a catalytic Lac1 subunit and a regulatory Lip1 subunit, in complex with C26-CoA substrate. The CerS holoenzyme exists as a dimer of Lac1-Lip1 heterodimers. Lac1 contains a hydrophilic reaction chamber and a hydrophobic tunnel for binding the CoA moiety and C26-acyl chain of C26-CoA, respectively. Lip1 interacts with both the transmembrane region and the last luminal loop of Lac1 to maintain the proper acyl chain binding tunnel. A lateral opening on Lac1 serves as a potential entrance for the sphingoid base substrate. Our findings provide a template for understanding the working mechanism of eukaryotic ceramide synthases and may facilitate the development of therapeutic CerS modulators.
History
DepositionApr 7, 2023-
Header (metadata) releaseDec 13, 2023-
Map releaseDec 13, 2023-
UpdateDec 20, 2023-
Current statusDec 20, 2023Processing site: PDBc / Status: Released

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Structure visualization

Supplemental images

emd_35862.png

Downloads & links

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Map

FileDownload / File: emd_35862.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.072 Å
Density
Contour LevelBy AUTHOR: 0.37
Minimum - Maximum-2.5765014 - 4.1975617
Average (Standard dev.)-0.0013432548 (±0.09184291)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 274.432 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #2

Fileemd_35862_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_35862_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 : Lac1-Lip1 complex

EntireName: Lac1-Lip1 complex
Components
  • Complex: Lac1-Lip1 complex
    • Protein or peptide: Ceramide synthase LAC1
    • Protein or peptide: Ceramide synthase subunit LIP1
  • Ligand: (4S,7R)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE
  • Ligand: Hexacosanoyl-CoA

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Supramolecule #1: Lac1-Lip1 complex

SupramoleculeName: Lac1-Lip1 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)

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Macromolecule #1: Ceramide synthase LAC1

MacromoleculeName: Ceramide synthase LAC1 / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: very-long-chain ceramide synthase
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Molecular weightTheoretical: 50.289082 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MSTIKPSPSN NNLKVRSRPR RKSSIGKIDL GDTVPSLGTM FETKESKTAA KRRMQRLSEA TKNDSDLVKK IWFSFREISY RHAWIAPLM ILIAVYSAYF TSGNTTKTNV LHRFVAVSYQ IGDTNAYGKG INDLCFVFYY MIFFTFLREF LMDVVIRPFA I RLHVTSKH ...String:
MSTIKPSPSN NNLKVRSRPR RKSSIGKIDL GDTVPSLGTM FETKESKTAA KRRMQRLSEA TKNDSDLVKK IWFSFREISY RHAWIAPLM ILIAVYSAYF TSGNTTKTNV LHRFVAVSYQ IGDTNAYGKG INDLCFVFYY MIFFTFLREF LMDVVIRPFA I RLHVTSKH RIKRIMEQMY AIFYTGVSGP FGIYCMYHSD LWFFNTKAMY RTYPDFTNPF LFKVFYLGQA AFWAQQACIL VL QLEKPRK DHNELTFHHI VTLLLIWSSY VFHFTKMGLP IYITMDVSDF LLSFSKTLNY LDSGLAFFSF AIFVVAWIYL RHY INLKIL WSVLTQFRTE GNYVLNFATQ QYKCWISLPI VFVLIGALQL VNLYWLFLIF RVLYRILWRG ILKDDRSDSE SDEE SDESS TTPTDSTPTK KDILEDYKDD DDK

UniProtKB: Ceramide synthase LAC1

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Macromolecule #2: Ceramide synthase subunit LIP1

MacromoleculeName: Ceramide synthase subunit LIP1 / type: protein_or_peptide / ID: 2 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Molecular weightTheoretical: 17.228682 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
MSQPTPIITT KSAAKPKPKI FNLFRVCFIS LLLIAAVEYF KYGTRINYEW FHCTPIKEPQ SGSVIKLWAR GGPSCDKRGE YKTIVKRIT RDYEPNDEHL SFCIIENDNV PPVHYPIHED KGEPGYVAYV GYDTDSELVQ ELCADSTIYH M

UniProtKB: Ceramide synthase subunit LIP1

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Macromolecule #3: (4S,7R)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-...

MacromoleculeName: (4S,7R)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE
type: ligand / ID: 3 / Number of copies: 6 / Formula: 6PL
Molecular weightTheoretical: 763.1 Da
Chemical component information

ChemComp-6PL:
(4S,7R)-4-HYDROXY-N,N,N-TRIMETHYL-9-OXO-7-[(PALMITOYLOXY)METHYL]-3,5,8-TRIOXA-4-PHOSPHAHEXACOSAN-1-AMINIUM 4-OXIDE / phospholipid*YM

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Macromolecule #4: Hexacosanoyl-CoA

MacromoleculeName: Hexacosanoyl-CoA / type: ligand / ID: 4 / Number of copies: 2 / Formula: 9NY
Molecular weightTheoretical: 1.146209 KDa
Chemical component information

ChemComp-9NY:
Hexacosanoyl-CoA

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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: 7
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Initial angle assignmentType: ANGULAR RECONSTITUTION
Final angle assignmentType: ANGULAR RECONSTITUTION
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.09 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 179070

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