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- EMDB-30074: cryo-EM structure of Scap/Insig complex in the present of 25-hydr... -

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Basic information

Entry
Database: EMDB / ID: EMD-30074
Titlecryo-EM structure of Scap/Insig complex in the present of 25-hydroxyl cholesterol.
Map datacryo-EM structure of Scap/Insig in the present of 25HC
Sample
  • Complex: Scap and Insig complex
    • Protein or peptide: Insulin-induced gene 2 protein
    • Protein or peptide: Sterol regulatory element-binding protein cleavage-activating protein,Sterol regulatory element-binding protein cleavage-activating protein
  • Ligand: 25-HYDROXYCHOLESTEROL
KeywordsScap / Insig / cholesterol / sterol sensing / 25-hydroxycholesterol / SREBP / MEMBRANE PROTEIN
Function / homology
Function and homology information


SREBP-SCAP complex retention in endoplasmic reticulum / SREBP-SCAP-Insig complex / cranial suture morphogenesis / SREBP-SCAP complex / negative regulation of steroid biosynthetic process / regulation of cholesterol biosynthetic process / sterol binding / SREBP signaling pathway / COPII-coated vesicle cargo loading / negative regulation of cholesterol biosynthetic process ...SREBP-SCAP complex retention in endoplasmic reticulum / SREBP-SCAP-Insig complex / cranial suture morphogenesis / SREBP-SCAP complex / negative regulation of steroid biosynthetic process / regulation of cholesterol biosynthetic process / sterol binding / SREBP signaling pathway / COPII-coated vesicle cargo loading / negative regulation of cholesterol biosynthetic process / regulation of fatty acid biosynthetic process / negative regulation of fatty acid biosynthetic process / Regulation of cholesterol biosynthesis by SREBP (SREBF) / oxysterol binding / response to vitamin B3 / middle ear morphogenesis / inner ear morphogenesis / triglyceride metabolic process / cholesterol biosynthetic process / roof of mouth development / cholesterol metabolic process / protein sequestering activity / response to insulin / positive regulation of cholesterol biosynthetic process / ER to Golgi transport vesicle membrane / cellular response to insulin stimulus / unfolded protein binding / response to hypoxia / immune response / Golgi membrane / endoplasmic reticulum membrane / protein-containing complex binding / endoplasmic reticulum / Golgi apparatus / membrane
Similarity search - Function
Insulin-induced protein family / Insulin-induced protein (INSIG) / SCAP N-terminal / SCAP Beta-propeller / Sterol regulatory element-binding protein cleavage-activating protein / : / Sterol-sensing domain of SREBP cleavage-activation / Sterol-sensing domain (SSD) profile. / Sterol-sensing domain / WD40 repeat, conserved site ...Insulin-induced protein family / Insulin-induced protein (INSIG) / SCAP N-terminal / SCAP Beta-propeller / Sterol regulatory element-binding protein cleavage-activating protein / : / Sterol-sensing domain of SREBP cleavage-activation / Sterol-sensing domain (SSD) profile. / Sterol-sensing domain / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Sterol regulatory element-binding protein cleavage-activating protein / Insulin-induced gene 2 protein
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsYan R / Cao P
Funding support China, 1 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China) China
CitationJournal: Science / Year: 2021
Title: A structure of human Scap bound to Insig-2 suggests how their interaction is regulated by sterols.
Authors: Renhong Yan / Pingping Cao / Wenqi Song / Hongwu Qian / Ximing Du / Hudson W Coates / Xin Zhao / Yaning Li / Shuai Gao / Xin Gong / Ximing Liu / Jianhua Sui / Jianlin Lei / Hongyuan Yang / ...Authors: Renhong Yan / Pingping Cao / Wenqi Song / Hongwu Qian / Ximing Du / Hudson W Coates / Xin Zhao / Yaning Li / Shuai Gao / Xin Gong / Ximing Liu / Jianhua Sui / Jianlin Lei / Hongyuan Yang / Andrew J Brown / Qiang Zhou / Chuangye Yan / Nieng Yan /
Abstract: The sterol regulatory element-binding protein (SREBP) pathway controls cellular homeostasis of sterols. The key players in this pathway, Scap and Insig-1 and -2, are membrane-embedded sterol sensors. ...The sterol regulatory element-binding protein (SREBP) pathway controls cellular homeostasis of sterols. The key players in this pathway, Scap and Insig-1 and -2, are membrane-embedded sterol sensors. The 25-hydroxycholesterol (25HC)-dependent association of Scap and Insig acts as the master switch for the SREBP pathway. Here, we present cryo-electron microscopy analysis of the human Scap and Insig-2 complex in the presence of 25HC, with the transmembrane (TM) domains determined at an average resolution of 3.7 angstrom. The sterol-sensing domain in Scap and all six TMs in Insig-2 were resolved. A 25HC molecule is sandwiched between the S4 to S6 segments in Scap and TMs 3 and 4 in Insig-2 in the luminal leaflet of the membrane. Unwinding of the middle of the Scap-S4 segment is crucial for 25HC binding and Insig association.
History
DepositionMar 6, 2020-
Header (metadata) releaseJan 20, 2021-
Map releaseJan 20, 2021-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.85
  • Imaged by UCSF Chimera
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  • Surface view colored by height
  • Surface level: 0.85
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6m49
  • Surface level: 0.85
  • Imaged by UCSF Chimera
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  • Simplified surface model + fitted atomic model
  • Atomic modelsPDB-6m49
  • Imaged by Jmol
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_30074.png

Downloads & links

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Map

FileDownload / File: emd_30074.map.gz / Format: CCP4 / Size: 30.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationcryo-EM structure of Scap/Insig in the present of 25HC
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.09 Å/pix.
x 200 pix.
= 217.4 Å		1.09 Å/pix.
x 200 pix.
= 217.4 Å		1.09 Å/pix.
x 200 pix.
= 217.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.087 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.85 / Movie #1: 0.85
Minimum - Maximum-2.7098722 - 3.9443238
Average (Standard dev.)0.0070932517 (±0.10552889)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions200200200
Spacing200200200
CellA=B=C: 217.40001 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0871.0871.087
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z217.400217.400217.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-2.7103.9440.007

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Supplemental data

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Additional map: #1

Fileemd_30074_additional_1.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 : Scap and Insig complex

EntireName: Scap and Insig complex
Components
  • Complex: Scap and Insig complex
    • Protein or peptide: Insulin-induced gene 2 protein
    • Protein or peptide: Sterol regulatory element-binding protein cleavage-activating protein,Sterol regulatory element-binding protein cleavage-activating protein
  • Ligand: 25-HYDROXYCHOLESTEROL

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Supramolecule #1: Scap and Insig complex

SupramoleculeName: Scap and Insig complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#2
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Insulin-induced gene 2 protein

MacromoleculeName: Insulin-induced gene 2 protein / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 24.74966 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MAEGETESPG PKKSGPYISS VTSQSVNLMI RGVVLFFIGV FLALVLNLLQ IQRNVTLFPP DVIASIFSSA WWVPPCCGTA SAVIGLLYP SIDRHLGEPH KFKREWSSVM RCVAVFVGIN HASAKVDFDN NIQLSLTLAA LSIGLWWTFD RSRSGFGLGV G IAFLATVV ...String:
MAEGETESPG PKKSGPYISS VTSQSVNLMI RGVVLFFIGV FLALVLNLLQ IQRNVTLFPP DVIASIFSSA WWVPPCCGTA SAVIGLLYP SIDRHLGEPH KFKREWSSVM RCVAVFVGIN HASAKVDFDN NIQLSLTLAA LSIGLWWTFD RSRSGFGLGV G IAFLATVV TQLLVYNGVY QYTSPDFLYV RSWLPCIFFA GGITMGNIGR QLAMYESKVI AEKSHQE

UniProtKB: Insulin-induced gene 2 protein

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Macromolecule #2: Sterol regulatory element-binding protein cleavage-activating pro...

MacromoleculeName: Sterol regulatory element-binding protein cleavage-activating protein,Sterol regulatory element-binding protein cleavage-activating protein
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 63.295406 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: MTLTERLREK ISRAFYNHGL LCASYPIPII LFTGFCILAC CYPLLKLPLP GTVPGKYSGV SLYTRKRMVS YTITLVFQHY HAKFLGSLR ARLMLLHPSP NCSLRAESLV HVHFKEEIGV AELIPLVTTY IILFAYIYFS TRKIDMVKSK WGLALAAVVT V LSSLLMSV ...String:
MTLTERLREK ISRAFYNHGL LCASYPIPII LFTGFCILAC CYPLLKLPLP GTVPGKYSGV SLYTRKRMVS YTITLVFQHY HAKFLGSLR ARLMLLHPSP NCSLRAESLV HVHFKEEIGV AELIPLVTTY IILFAYIYFS TRKIDMVKSK WGLALAAVVT V LSSLLMSV GLCTLFGLTP TLNGGEIFPY LVVVIGLENV LVLTKSVVST PVDLEVKLRI AQGLSSESWS IMKNMATELG II LIGYFTL VPAIQEFCLF AVVGLVSDFF LQMLFFTTVL SIDIRRMELA DLNKRLPPEA CLPSAKPVGQ PTRYERQLAV RPS TPHTIT LQPSSFRNLR LPKRLRVVYF LARTRLAQRL IMAGTVVWIG ILVYTDPAGL RNYLAAQVTE QSPLGEGALA PMPV PSGML PPSHPDPAFS IFPPDAPKLP ENQTSPGESP ERGGPAEVVH DSPVPEVTWG PEDEELWRKL SFRHWPTLFS YYNIT LAKR YISLLPVIPV TLRLNPREAL EGRHPQDGRS AWPPPGPIPA GHWEAGPKGP GGVQAHGDVT LYKVAALGLA TGIVLV LLL LCLYRVLCP

UniProtKB: Sterol regulatory element-binding protein cleavage-activating protein, Sterol regulatory element-binding protein cleavage-activating protein

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Macromolecule #3: 25-HYDROXYCHOLESTEROL

MacromoleculeName: 25-HYDROXYCHOLESTEROL / type: ligand / ID: 3 / Number of copies: 1 / Formula: HC3
Molecular weightTheoretical: 402.653 Da
Chemical component information

ChemComp-HC3:
25-HYDROXYCHOLESTEROL

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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

Concentration10 mg/mL
BufferpH: 8
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: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
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: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 153168
Initial angle assignmentType: PROJECTION MATCHING
Final angle assignmentType: PROJECTION MATCHING

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