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- PDB-7yjm: Cryo-EM structure of the monomeric atSPT-ORM1 complex -

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Basic information

Entry
Database: PDB / ID: 7yjm
TitleCryo-EM structure of the monomeric atSPT-ORM1 complex
Components
  • (atLCB1) x 2
  • Long chain base biosynthesis protein 2a
  • ORMDL family protein
  • Transmembrane protein, putative (DUF3317)Transmembrane protein
KeywordsTRANSFERASE/INHIBITOR / ceramide / TRANSFERASE-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
PYRIDOXAL-5'-PHOSPHATE / Chem-Z1T / 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)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.2 Å
AuthorsXie, T. / Liu, P. / Gong, X.
Funding support1items
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, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Apr 5, 2023Provider: repository / Type: Initial release
Revision 1.1Apr 12, 2023Group: Database references / Category: citation / Item: _citation.pdbx_database_id_PubMed
Revision 1.2Apr 19, 2023Group: Database references / Category: citation / citation_author
Item: _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID
Revision 1.3Mar 20, 2024Group: Data collection / Source and taxonomy / Category: chem_comp_atom / chem_comp_bond / entity_src_gen

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: atLCB1
B: Long chain base biosynthesis protein 2a
D: ORMDL family protein
E: atLCB1
C: Transmembrane protein, putative (DUF3317)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)136,1457
Polymers135,2485
Non-polymers8972
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 5 types, 5 molecules ABDEC

#1: Protein atLCB1


Mass: 46276.117 Da / Num. of mol.: 1 / Fragment: UNP residues 63-482
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: Q94IB8
#2: Protein Long chain base biosynthesis protein 2a / AtLCB2a / AtLCB2


Mass: 54359.457 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: LCB2a, LCB2 / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: Q9LSZ9, serine C-palmitoyltransferase
#3: Protein ORMDL family protein / atORM1


Mass: 18220.266 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: Q9C5I0
#4: Protein atLCB1


Mass: 6949.384 Da / Num. of mol.: 1 / Fragment: UNP residues 1-62
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: LCB1, EMB2779, FBR11, At4g36480, AP22, C7A10.880 / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: Q94IB8, serine C-palmitoyltransferase
#5: Protein Transmembrane protein, putative (DUF3317) / Transmembrane protein / atSPTssa


Mass: 9442.687 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Arabidopsis thaliana (thale cress) / Gene: At1g06515 / Cell line (production host): HEK293 / Production host: Homo sapiens (human) / References: UniProt: A8MSB8

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Non-polymers , 2 types, 2 molecules

#6: Chemical ChemComp-PLP / PYRIDOXAL-5'-PHOSPHATE / VITAMIN B6 Phosphate / Pyridoxal phosphate


Mass: 247.142 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Formula: C8H10NO6P
#7: Chemical ChemComp-Z1T / N-[(2S,3R,4E)-1,3-dihydroxyoctadec-4-en-2-yl]tetracosanamide


Mass: 650.113 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C42H83NO3 / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: SPT-ORM1 complex / Type: COMPLEX / Entity ID: #1-#3, #5 / Source: RECOMBINANT
Source (natural)Organism: Arabidopsis thaliana (thale cress)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2000 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.18.1_3865: / Classification: refinement
CTF correctionType: NONE
3D reconstructionResolution: 3.2 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 115440 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0059166
ELECTRON MICROSCOPYf_angle_d0.65712422
ELECTRON MICROSCOPYf_dihedral_angle_d30.7131236
ELECTRON MICROSCOPYf_chiral_restr0.0471389
ELECTRON MICROSCOPYf_plane_restr0.0051564

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