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- PDB-8erp: Structure of Xenopus cholinephosphotransferase1 in complex with C... -

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Basic information

Entry
Database: PDB / ID: 8erp
TitleStructure of Xenopus cholinephosphotransferase1 in complex with CDP-choline
ComponentsCholinephosphotransferase 1
KeywordsTRANSFERASE / CDP-APs / phospholipid synthesis
Function / homology
Function and homology information


diacylglycerol cholinephosphotransferase / diacylglycerol cholinephosphotransferase activity / ethanolaminephosphotransferase activity / phosphatidylethanolamine biosynthetic process / Golgi membrane / endoplasmic reticulum membrane / metal ion binding
Similarity search - Function
Choline/ethanolamine phosphotransferase / : / CDP-alcohol phosphatidyltransferase / CDP-alcohol phosphatidyltransferase, transmembrane domain / CDP-alcohol phosphatidyltransferase / CDP-alcohol phosphatidyltransferases signature.
Similarity search - Domain/homology
Chem-CDC / 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine / Cholinephosphotransferase 1
Similarity search - Component
Biological speciesXenopus laevis (African clawed frog)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsWang, L. / Zhou, M.
Funding support United States, 3items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)DK122784 United States
National Institutes of Health/National Heart, Lung, and Blood Institute (NIH/NHLBI)HL086392 United States
Cancer Prevention and Research Institute of Texas (CPRIT)R1223 United States
CitationJournal: Nat Commun / Year: 2023
Title: Structure of a eukaryotic cholinephosphotransferase-1 reveals mechanisms of substrate recognition and catalysis.
Authors: Lie Wang / Ming Zhou /
Abstract: Phosphatidylcholine (PC) is the most abundant phospholipid in eukaryotic cell membranes. In eukaryotes, two highly homologous enzymes, cholinephosphotransferase-1 (CHPT1) and choline/ethanolamine ...Phosphatidylcholine (PC) is the most abundant phospholipid in eukaryotic cell membranes. In eukaryotes, two highly homologous enzymes, cholinephosphotransferase-1 (CHPT1) and choline/ethanolamine phosphotransferase-1 (CEPT1) catalyze the final step of de novo PC synthesis. CHPT1/CEPT1 joins two substrates, cytidine diphosphate-choline (CDP-choline) and diacylglycerol (DAG), to produce PC, and Mg is required for the reaction. However, mechanisms of substrate recognition and catalysis remain unresolved. Here we report structures of a CHPT1 from Xenopus laevis (xlCHPT1) determined by cryo-electron microscopy to an overall resolution of ~3.2 Å. xlCHPT1 forms a homodimer, and each protomer has 10 transmembrane helices (TMs). The first 6 TMs carve out a cone-shaped enclosure in the membrane in which the catalysis occurs. The enclosure opens to the cytosolic side, where a CDP-choline and two Mg are coordinated. The structures identify a catalytic site unique to eukaryotic CHPT1/CEPT1 and suggest an entryway for DAG. The structures also reveal an internal pseudo two-fold symmetry between TM3-6 and TM7-10, and suggest that CHPT1/CEPT1 may have evolved from their distant prokaryotic ancestors through gene duplication.
History
DepositionOct 12, 2022Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 26, 2023Provider: repository / Type: Initial release
Revision 1.1May 3, 2023Group: Database references / Structure summary / Category: audit_author / citation_author / Item: _audit_author.name / _citation_author.name
Revision 1.2Nov 8, 2023Group: Data collection / Database references
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
B: Cholinephosphotransferase 1
A: Cholinephosphotransferase 1
hetero molecules


Theoretical massNumber of molelcules
Total (without water)105,36028
Polymers89,0842
Non-polymers16,27526
Water0
1


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

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Components

#1: Protein Cholinephosphotransferase 1 / Diacylglycerol cholinephosphotransferase 1


Mass: 44542.230 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Xenopus laevis (African clawed frog) / Gene: chpt1 / Production host: Spodoptera frugiperda (fall armyworm)
References: UniProt: Q4KLV1, diacylglycerol cholinephosphotransferase
#2: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Mg
#3: Chemical
ChemComp-LBN / 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine / (2R)-2-[(9Z)-9-Octadecenoyloxy]-3-(palmitoyloxy)propyl 2-(trimethylammonio)ethyl phosphate / POPC


Mass: 760.076 Da / Num. of mol.: 20 / Source method: obtained synthetically / Formula: C42H82NO8P / Comment: phospholipid*YM
#4: Chemical ChemComp-CDC / [2-CYTIDYLATE-O'-PHOSPHONYLOXYL]-ETHYL-TRIMETHYL-AMMONIUM / Citicoline


Mass: 488.324 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C14H26N4O11P2 / Feature type: SUBJECT OF INVESTIGATION
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: homodimer of choline phosphotransferase 1 / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Source (natural)Organism: Xenopus laevis (African clawed frog)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
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: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: SPOT SCAN
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 20000 nm / Nominal defocus min: 8000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.1_4122: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 381720 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0036202
ELECTRON MICROSCOPYf_angle_d0.6528344
ELECTRON MICROSCOPYf_dihedral_angle_d5.8991074
ELECTRON MICROSCOPYf_chiral_restr0.042956
ELECTRON MICROSCOPYf_plane_restr0.006982

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