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- EMDB-33875: Cryo-EM structure of the monomeric atSPT-ORM1 (ORM1-N17A) complex -

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Basic information

Entry
Database: EMDB / ID: EMD-33875
TitleCryo-EM structure of the monomeric atSPT-ORM1 (ORM1-N17A) complex
Map data
Sample
  • Complex: SPT-ORM1 complex
    • Protein or peptide: Long chain base biosynthesis protein 1
    • Protein or peptide: Long chain base biosynthesis protein 2a
    • Protein or peptide: ORMDL family protein
    • Protein or peptide: Transmembrane protein, putative (DUF3317)Transmembrane protein
  • Protein or peptide: Long chain base biosynthesis protein 1
  • Ligand: PYRIDOXAL-5'-PHOSPHATEPyridoxal phosphate
KeywordsTRANSFERASE-INHIBITOR complex
Function / homology
Function and homology information


positive regulation of serine C-palmitoyltransferase activity / regulation of sphingolipid biosynthetic process / multidimensional cell growth / leaf senescence / intracellular sphingolipid homeostasis / photomorphogenesis / serine C-palmitoyltransferase activity / pollen development / serine C-palmitoyltransferase / sphingosine biosynthetic process ...positive regulation of serine C-palmitoyltransferase activity / regulation of sphingolipid biosynthetic process / multidimensional cell growth / leaf senescence / intracellular sphingolipid homeostasis / photomorphogenesis / serine C-palmitoyltransferase activity / pollen development / serine C-palmitoyltransferase / sphingosine biosynthetic process / regulation of programmed cell death / embryo development ending in seed dormancy / sphingolipid biosynthetic process / vacuole / response to endoplasmic reticulum stress / pyridoxal phosphate binding / response to oxidative stress / defense response to bacterium / protein-containing complex binding / endoplasmic reticulum membrane / endoplasmic reticulum / membrane
Similarity search - Function
Small subunit of serine palmitoyltransferase-like / Small subunit of serine palmitoyltransferase-like / ORMDL family / ORMDL family / Aminotransferase, class-II, pyridoxal-phosphate binding site / Aminotransferases class-II pyridoxal-phosphate attachment site. / Aminotransferase, class I/classII / Aminotransferase class I and II / Pyridoxal phosphate-dependent transferase, small domain / Pyridoxal phosphate-dependent transferase, major domain / Pyridoxal phosphate-dependent transferase
Similarity search - Domain/homology
Transmembrane protein, putative (DUF3317) / Long chain base biosynthesis protein 1 / ORMDL family protein / Long chain base biosynthesis protein 2a
Similarity search - Component
Biological speciesArabidopsis thaliana (thale cress)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsXie T / Liu P / Gong X
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Sci Adv / Year: 2023
Title: Mechanism of sphingolipid homeostasis revealed by structural analysis of SPT-ORM1 complex.
Authors: Peng Liu / Tian Xie / Xinyue Wu / Gongshe Han / Sita D Gupta / Zike Zhang / Jian Yue / Feitong Dong / Kenneth Gable / Somashekarappa Niranjanakumari / Wanyuan Li / Lin Wang / Wenchen Liu / ...Authors: Peng Liu / Tian Xie / Xinyue Wu / Gongshe Han / Sita D Gupta / Zike Zhang / Jian Yue / Feitong Dong / Kenneth Gable / Somashekarappa Niranjanakumari / Wanyuan Li / Lin Wang / Wenchen Liu / Ruifeng Yao / Edgar B Cahoon / Teresa M Dunn / Xin Gong /
Abstract: The serine palmitoyltransferase (SPT) complex catalyzes the first and rate-limiting step in sphingolipid biosynthesis in all eukaryotes. ORM/ORMDL proteins are negative regulators of SPT that respond ...The serine palmitoyltransferase (SPT) complex catalyzes the first and rate-limiting step in sphingolipid biosynthesis in all eukaryotes. ORM/ORMDL proteins are negative regulators of SPT that respond to cellular sphingolipid levels. However, the molecular basis underlying ORM/ORMDL-dependent homeostatic regulation of SPT is not well understood. We determined the cryo-electron microscopy structure of SPT-ORM1 complex, composed of LCB1, LCB2a, SPTssa, and ORM1, in an inhibited state. A ceramide molecule is sandwiched between ORM1 and LCB2a in the cytosolic membrane leaflet. Ceramide binding is critical for the ORM1-dependent SPT repression, and dihydroceramides and phytoceramides differentially affect this repression. A hybrid β sheet, formed by the amino termini of ORM1 and LCB2a and induced by ceramide binding, stabilizes the amino terminus of ORM1 in an inhibitory conformation. Our findings provide mechanistic insights into sphingolipid homeostatic regulation via the binding of ceramide to the SPT-ORM/ORMDL complex that may have implications for plant-specific processes such as the hypersensitive response for microbial pathogen resistance.
History
DepositionJul 20, 2022-
Header (metadata) releaseApr 5, 2023-
Map releaseApr 5, 2023-
UpdateMar 20, 2024-
Current statusMar 20, 2024Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_33875.png

Downloads & links

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Map

FileDownload / File: emd_33875.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-2.4545743 - 4.0853825
Average (Standard dev.)0.0011797562 (±0.10137654)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 276.48 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_33875_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_33875_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 : SPT-ORM1 complex

EntireName: SPT-ORM1 complex
Components
  • Complex: SPT-ORM1 complex
    • Protein or peptide: Long chain base biosynthesis protein 1
    • Protein or peptide: Long chain base biosynthesis protein 2a
    • Protein or peptide: ORMDL family protein
    • Protein or peptide: Transmembrane protein, putative (DUF3317)Transmembrane protein
  • Protein or peptide: Long chain base biosynthesis protein 1
  • Ligand: PYRIDOXAL-5'-PHOSPHATEPyridoxal phosphate

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Supramolecule #1: SPT-ORM1 complex

SupramoleculeName: SPT-ORM1 complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3, #5
Source (natural)Organism: Arabidopsis thaliana (thale cress)

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Macromolecule #1: Long chain base biosynthesis protein 1

MacromoleculeName: Long chain base biosynthesis protein 1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 46.276117 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: PLTEQEIDEL CDEWVPEPLI PPITEDMKHE PPVLESAAGP HTTVNGKDVV NFASANYLGL IGHEKLLESC TSALEKYGVG SCGPRGFYG TIDVHLDCET RISKFLGTPD SILYSYGLST MFSTIPCFCK KGDVIVADEG VHWGIQNGLQ LSRSTIVYFK H NDMESLRI ...String:
PLTEQEIDEL CDEWVPEPLI PPITEDMKHE PPVLESAAGP HTTVNGKDVV NFASANYLGL IGHEKLLESC TSALEKYGVG SCGPRGFYG TIDVHLDCET RISKFLGTPD SILYSYGLST MFSTIPCFCK KGDVIVADEG VHWGIQNGLQ LSRSTIVYFK H NDMESLRI TLEKIMTKYK RSKNLRRYIV AEAVYQNSGQ IAPLDEIVKL KEKYRFRVIL DESNSFGVLG RSGRGLAEHH SV PIEKIDV VTAAMGHALA TEGGFCTGNA RIIDYQRLSS SGYVFSASLP PYLASAAITA IDVIDQNPDM LVKLKQNVAL LWK GLSDIK GMSLTSNRES PIVFLKLEKS SGSAKDDLLL LEKMADRALK EDSLLVVSSK RSFLDKCRLP VGIKLYVSAG HSES DLLKA SESLKRLASE LLLKS

UniProtKB: Long chain base biosynthesis protein 1

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Macromolecule #2: Long chain base biosynthesis protein 2a

MacromoleculeName: Long chain base biosynthesis protein 2a / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO / EC number: serine C-palmitoyltransferase
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 54.359457 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MITIPYLTAV STYFSYGLLF AFGQLRDFFR RFIDWWFTSN LQGYAPICLG HEDFYIRRLY HRIQDCFERP ISSAPDAWFD VVERYSNDN NKTLKRTTKT SRCLNLGSYN YLGFGSFDEY CTPRVIESLK KFSASTCSSR VDAGTTSVHA ELEECVTRFV G KPAAVVFG ...String:
MITIPYLTAV STYFSYGLLF AFGQLRDFFR RFIDWWFTSN LQGYAPICLG HEDFYIRRLY HRIQDCFERP ISSAPDAWFD VVERYSNDN NKTLKRTTKT SRCLNLGSYN YLGFGSFDEY CTPRVIESLK KFSASTCSSR VDAGTTSVHA ELEECVTRFV G KPAAVVFG MGYATNSAII PVLIGKGGLI ISDSLNHSSI VNGARGSGAT IRVFQHNTPS HLERVLREQI AEGQPRTHRP WK KIIVVVE GIYSMEGEIC HLPEVVAICK KYKAYVYLDE AHSIGAIGKT GKGICELLGV DTADVDVMMG TFTKSFGSCG GYI AGSKEL IQYLKHQCPA HLYATSIPTP SAQQIISAIK VILGEDGSNR GAQKLARIRE NSNFFRAELQ KMGFEVLGDN DSPV MPIML YNPAKIPAFS RECLRQKVAV VVVGFPATPL LLARARICIS ASHSREDLIR ALKVISKVGD LSGIKYFPAE PKKIE QSKN DIKLD

UniProtKB: Long chain base biosynthesis protein 2a

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Macromolecule #3: ORMDL family protein

MacromoleculeName: ORMDL family protein / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 18.17724 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
MANLYVKAVP PPDMNRATEW FMYPGVWTTY MLILFFGWLV VLSVSGCSPG MAWTVVNLAH FVVTYHSFHW MKGTPFADDQ GIYNGLTWW EQMDNGQQLT RNRKFLTLVP VVLYLIASHT TDYRHPWLFL NTLAVMVLVV AKFPNMHKVR IFGINGDK

UniProtKB: ORMDL family protein

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Macromolecule #4: Long chain base biosynthesis protein 1

MacromoleculeName: Long chain base biosynthesis protein 1 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 6.949384 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
MASNLVEMFN AALNWVTMIL ESPSARVVLF GVPIRGHFFV EGLLGVVIII LLTRKSYKPP KR

UniProtKB: Long chain base biosynthesis protein 1

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Macromolecule #5: Transmembrane protein, putative (DUF3317)

MacromoleculeName: Transmembrane protein, putative (DUF3317) / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Molecular weightTheoretical: 9.442687 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString:
MADYKDDDDK SGPDEVDASG RMNWVQRKIY LYNVTFGLYM LDWWERYLFN SLVVVLMWFV LYNGTRYFSE LFQRHLT

UniProtKB: Transmembrane protein, putative (DUF3317)

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Macromolecule #6: PYRIDOXAL-5'-PHOSPHATE

MacromoleculeName: PYRIDOXAL-5'-PHOSPHATE / type: ligand / ID: 6 / Number of copies: 1 / Formula: PLP
Molecular weightTheoretical: 247.142 Da
Chemical component information

ChemComp-PLP:
PYRIDOXAL-5'-PHOSPHATE / Pyridoxal phosphate

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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: 7.5
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 K2 SUMMIT (4k 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: OTHER
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 58458

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