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- EMDB-26707: Human PORCN in complex with Palmitoleoyl-CoA -

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Basic information

Entry
Database: EMDB / ID: EMD-26707
TitleHuman PORCN in complex with Palmitoleoyl-CoA
Map datamembrane protein
Sample
  • Complex: 16:1 CoA-bound human PORCN
    • Complex: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine (E.C.2.3.1.250)
      • Protein or peptide: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine
    • Complex: 2C11 light chain, 2C11 heavy chain
      • Protein or peptide: 2C11 light chain
      • Protein or peptide: 2C11 heavy chain
  • Ligand: ~{S}-[2-[3-[[(2~{R})-4-[[[(2~{R},3~{R},4~{R},5~{R})-5-(6-aminopurin-9-yl)-4-oxidanyl-3-phosphonooxy-oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-3,3-dimethyl-2-oxidanyl-butanoyl]amino]propanoylamino]ethyl] (~{Z})-hexadec-9-enethioate
  • Ligand: ZINC ION
  • Ligand: CHOLESTEROL
  • Ligand: Digitonin
  • Ligand: OLEIC ACID
KeywordsCoA-bound / complex / TRANSFERASE / TRANSFERASE-TRANSFERASE INHIBITOR complex
Function / homology
Function and homology information


[Wnt protein] O-palmitoleoyl transferase / protein palmitoleylation / palmitoleoyltransferase activity / LGK974 inhibits PORCN / protein lipidation / Wnt protein secretion / lipid modification / glycoprotein metabolic process / WNT ligand biogenesis and trafficking / Wnt-protein binding ...[Wnt protein] O-palmitoleoyl transferase / protein palmitoleylation / palmitoleoyltransferase activity / LGK974 inhibits PORCN / protein lipidation / Wnt protein secretion / lipid modification / glycoprotein metabolic process / WNT ligand biogenesis and trafficking / Wnt-protein binding / AMPA glutamate receptor complex / regulation of postsynaptic membrane neurotransmitter receptor levels / Wnt signaling pathway / endoplasmic reticulum membrane / glutamatergic synapse / endoplasmic reticulum / membrane
Similarity search - Function
: / Membrane bound O-acyl transferase, MBOAT / MBOAT, membrane-bound O-acyltransferase family
Similarity search - Domain/homology
Protein-serine O-palmitoleoyltransferase porcupine
Similarity search - Component
Biological speciesHomo sapiens (human) / Mus musculus (house mouse)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.11 Å
AuthorsLiu Y / Qi X / Li X
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS) United States
Welch Foundation United States
CitationJournal: Nature / Year: 2022
Title: Mechanisms and inhibition of Porcupine-mediated Wnt acylation.
Authors: Yang Liu / Xiaofeng Qi / Linda Donnelly / Nadia Elghobashi-Meinhardt / Tao Long / Rich W Zhou / Yingyuan Sun / Boyuan Wang / Xiaochun Li /
Abstract: Wnt signalling is essential for regulation of embryonic development and adult tissue homeostasis, and aberrant Wnt signalling is frequently associated with cancers. Wnt signalling requires ...Wnt signalling is essential for regulation of embryonic development and adult tissue homeostasis, and aberrant Wnt signalling is frequently associated with cancers. Wnt signalling requires palmitoleoylation on a hairpin 2 motif by the endoplasmic reticulum-resident membrane-bound O-acyltransferase Porcupine (PORCN). This modification is indispensable for Wnt binding to its receptor Frizzled, which triggers signalling. Here we report four cryo-electron microscopy structures of human PORCN: the complex with the palmitoleoyl-coenzyme A (palmitoleoyl-CoA) substrate; the complex with the PORCN inhibitor LGK974, an anti-cancer drug currently in clinical trials; the complex with LGK974 and WNT3A hairpin 2 (WNT3Ap); and the complex with a synthetic palmitoleoylated WNT3Ap analogue. The structures reveal that hairpin 2 of WNT3A, which is well conserved in all Wnt ligands, inserts into PORCN from the lumenal side, and the palmitoleoyl-CoA accesses the enzyme from the cytosolic side. The catalytic histidine triggers the transfer of the unsaturated palmitoleoyl group to the target serine on the Wnt hairpin 2, facilitated by the proximity of the two substrates. The inhibitor-bound structure shows that LGK974 occupies the palmitoleoyl-CoA binding site to prevent the reaction. Thus, this work provides a mechanism for Wnt acylation and advances the development of PORCN inhibitors for cancer treatment.
History
DepositionApr 21, 2022-
Header (metadata) releaseJul 20, 2022-
Map releaseJul 20, 2022-
UpdateNov 13, 2024-
Current statusNov 13, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_26707.png

Downloads & links

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Map

FileDownload / File: emd_26707.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationmembrane protein
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.83 Å/pix.
x 280 pix.
= 232.4 Å		0.83 Å/pix.
x 280 pix.
= 232.4 Å		0.83 Å/pix.
x 280 pix.
= 232.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.83 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.7
Minimum - Maximum-3.5774918 - 5.151635
Average (Standard dev.)0.0067899143 (±0.121688634)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 232.4 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: membrane protein

Fileemd_26707_half_map_1.map
Annotationmembrane protein
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: membrane protein

Fileemd_26707_half_map_2.map
Annotationmembrane protein
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 16:1 CoA-bound human PORCN

EntireName: 16:1 CoA-bound human PORCN
Components
  • Complex: 16:1 CoA-bound human PORCN
    • Complex: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine (E.C.2.3.1.250)
      • Protein or peptide: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine
    • Complex: 2C11 light chain, 2C11 heavy chain
      • Protein or peptide: 2C11 light chain
      • Protein or peptide: 2C11 heavy chain
  • Ligand: ~{S}-[2-[3-[[(2~{R})-4-[[[(2~{R},3~{R},4~{R},5~{R})-5-(6-aminopurin-9-yl)-4-oxidanyl-3-phosphonooxy-oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-3,3-dimethyl-2-oxidanyl-butanoyl]amino]propanoylamino]ethyl] (~{Z})-hexadec-9-enethioate
  • Ligand: ZINC ION
  • Ligand: CHOLESTEROL
  • Ligand: Digitonin
  • Ligand: OLEIC ACID

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Supramolecule #1: 16:1 CoA-bound human PORCN

SupramoleculeName: 16:1 CoA-bound human PORCN / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#3

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Supramolecule #2: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine (...

SupramoleculeName: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine (E.C.2.3.1.250)
type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: 2C11 light chain, 2C11 heavy chain

SupramoleculeName: 2C11 light chain, 2C11 heavy chain / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2-#3
Source (natural)Organism: Mus musculus (house mouse)

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Macromolecule #1: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine

MacromoleculeName: Isoform 2 of Protein-serine O-palmitoleoyltransferase porcupine
type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO / EC number: [Wnt protein] O-palmitoleoyl transferase
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 52.824652 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MDYKDDDDKA TFSRQEFFQQ LLQGCLLPTA QQGLDQIWLL LAICLACRLL WRLGLPSYLK HASTVAGGFF SLYHFFQLHM VWVVLLSLL CYLVLFLCRH SSHRGVFLSV TILIYLLMGE MHMVDTVTWH KMRGAQMIVA MKAVSLGFDL DRGEVGTVPS P VEFMGYLY ...String:
MDYKDDDDKA TFSRQEFFQQ LLQGCLLPTA QQGLDQIWLL LAICLACRLL WRLGLPSYLK HASTVAGGFF SLYHFFQLHM VWVVLLSLL CYLVLFLCRH SSHRGVFLSV TILIYLLMGE MHMVDTVTWH KMRGAQMIVA MKAVSLGFDL DRGEVGTVPS P VEFMGYLY FVGTIVFGPW ISFHSYLQAV QGRPLSCRWL QKVARSLALA LLCLVLSTCV GPYLFPYFIP LNGDRLLRNK KR KARWLRA YESAVSFHFS NYFVGFLSEA TATLAGAGFT EEKDHLEWDL TVSKPLNVEL PRSMVEVVTS WNLPMSYWLN NYV FKNALR LGTFSAVLVT YAASALLHGF SFHLAAVLLS LAFITYVEHV LRKRLARILS ACVLSKRCPP DCSHQHRLGL GVRA LNLLF GALAIFHLAY LGSLFDVDVD DTTEEQGYGM AYTVHKWSEL SWASHWVTFG CWIFYRLIG

UniProtKB: Protein-serine O-palmitoleoyltransferase porcupine

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Macromolecule #2: 2C11 light chain

MacromoleculeName: 2C11 light chain / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 25.673693 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGWSCIILFL VATARTGVHS DIHMTQSPAS LSAFVGETVT ITCRTSENIF SYLAWYQQKQ GKSPQLLVYN AKTLTSGVPS RFSGSGSGT QFSLKINSLQ PEDFGSYYCQ HHYGSPYTFG GGTKLEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF Y PREAKVQW ...String:
MGWSCIILFL VATARTGVHS DIHMTQSPAS LSAFVGETVT ITCRTSENIF SYLAWYQQKQ GKSPQLLVYN AKTLTSGVPS RFSGSGSGT QFSLKINSLQ PEDFGSYYCQ HHYGSPYTFG GGTKLEIKRT VAAPSVFIFP PSDEQLKSGT ASVVCLLNNF Y PREAKVQW KVDNALQSGN SQESVTEQDS KDSTYSLSST LTLSKADYEK HKVYACEVTH QGLSSPVTKS FNRGEC

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Macromolecule #3: 2C11 heavy chain

MacromoleculeName: 2C11 heavy chain / type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Mus musculus (house mouse)
Molecular weightTheoretical: 26.933135 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MGWSCIILFL VATATGVHSE IQLQQSGAEL VKPGASVKMS CKVSGYSFTG YNMNWVKQSH GKSLEWIGNI NPYYVSTNYN QKFTGKATF TVDRSSSTAY MQLDSLTSED SAVYYCARSY GSSHTFAYWG QGTLVTVSSA STKGPSVFPL APSSKSTSGG T AALGCLVK ...String:
MGWSCIILFL VATATGVHSE IQLQQSGAEL VKPGASVKMS CKVSGYSFTG YNMNWVKQSH GKSLEWIGNI NPYYVSTNYN QKFTGKATF TVDRSSSTAY MQLDSLTSED SAVYYCARSY GSSHTFAYWG QGTLVTVSSA STKGPSVFPL APSSKSTSGG T AALGCLVK DYFPEPVTVS WNSGALTSGV HTFPAVLQSS GLYSLSSVVT VPSSSLGTQT YICNVNHKPS NTKVDKRVEP KS CDKTHHH HHH

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Macromolecule #4: ~{S}-[2-[3-[[(2~{R})-4-[[[(2~{R},3~{R},4~{R},5~{R})-5-(6-aminopur...

MacromoleculeName: ~{S}-[2-[3-[[(2~{R})-4-[[[(2~{R},3~{R},4~{R},5~{R})-5-(6-aminopurin-9-yl)-4-oxidanyl-3-phosphonooxy-oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-3,3-dimethyl-2-oxidanyl- ...Name: ~{S}-[2-[3-[[(2~{R})-4-[[[(2~{R},3~{R},4~{R},5~{R})-5-(6-aminopurin-9-yl)-4-oxidanyl-3-phosphonooxy-oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-3,3-dimethyl-2-oxidanyl-butanoyl]amino]propanoylamino]ethyl] (~{Z})-hexadec-9-enethioate
type: ligand / ID: 4 / Number of copies: 1 / Formula: OH6
Molecular weightTheoretical: 1.003927 KDa
Chemical component information

ChemComp-OH6:
~{S}-[2-[3-[[(2~{R})-4-[[[(2~{R},3~{R},4~{R},5~{R})-5-(6-aminopurin-9-yl)-4-oxidanyl-3-phosphonooxy-oxolan-2-yl]methoxy-oxidanyl-phosphoryl]oxy-oxidanyl-phosphoryl]oxy-3,3-dimethyl-2-oxidanyl-butanoyl]amino]propanoylamino]ethyl] (~{Z})-hexadec-9-enethioate

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Macromolecule #5: ZINC ION

MacromoleculeName: ZINC ION / type: ligand / ID: 5 / Number of copies: 1 / Formula: ZN
Molecular weightTheoretical: 65.409 Da

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Macromolecule #6: CHOLESTEROL

MacromoleculeName: CHOLESTEROL / type: ligand / ID: 6 / Number of copies: 2 / Formula: CLR
Molecular weightTheoretical: 386.654 Da
Chemical component information

ChemComp-CLR:
CHOLESTEROL

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Macromolecule #7: Digitonin

MacromoleculeName: Digitonin / type: ligand / ID: 7 / Number of copies: 1 / Formula: AJP
Molecular weightTheoretical: 1.229312 KDa
Chemical component information

ChemComp-AJP:
Digitonin / detergent*YM

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Macromolecule #8: OLEIC ACID

MacromoleculeName: OLEIC ACID / type: ligand / ID: 8 / Number of copies: 1 / Formula: OLA
Molecular weightTheoretical: 282.461 Da
Chemical component information

ChemComp-OLA:
OLEIC ACID

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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
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 8.5 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.11 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 233902
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: NOT APPLICABLE

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