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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 data
SampleScap and Insig complex
  • Insulin-induced gene 2 protein
  • Sterol regulatory element-binding protein cleavage-activating protein,Sterol regulatory element-binding protein cleavage-activating protein
  • ligand
Function / homology
Function and homology information


SREBP-SCAP complex retention in endoplasmic reticulum / SREBP-SCAP-Insig complex / go:0045716: / SREBP signaling pathway / regulation of fatty acid biosynthetic process / sterol binding / COPII-coated vesicle cargo loading / cellular lipid metabolic process / sterol biosynthetic process / positive regulation of cholesterol biosynthetic process ...SREBP-SCAP complex retention in endoplasmic reticulum / SREBP-SCAP-Insig complex / go:0045716: / SREBP signaling pathway / regulation of fatty acid biosynthetic process / sterol binding / COPII-coated vesicle cargo loading / cellular lipid metabolic process / sterol biosynthetic process / positive regulation of cholesterol biosynthetic process / oxysterol binding / negative regulation of cholesterol biosynthetic process / cholesterol biosynthetic process / response to insulin / cholesterol metabolic process / ER to Golgi transport vesicle membrane / cellular response to insulin stimulus / unfolded protein binding / response to hypoxia / aging / immune response / Golgi membrane / endoplasmic reticulum membrane / protein-containing complex binding / Golgi apparatus / endoplasmic reticulum / protein-containing complex / integral component of membrane
Sterol-sensing domain / WD40 repeat, conserved site / WD40-repeat-containing domain superfamily / Sterol regulatory element-binding protein cleavage-activating protein / Insulin-induced protein family / WD40/YVTN repeat-like-containing domain superfamily / WD40-repeat-containing domain / WD40 repeat
Sterol regulatory element-binding protein cleavage-activating protein / Insulin-induced gene 2 protein
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsYan R / Cao P / Song W / Qian H / Du X / Coates HW / Zhao X / Li Y / Gao S / Gong X ...Yan R / Cao P / Song W / Qian H / Du X / Coates HW / Zhao X / Li Y / Gao S / Gong X / Liu X / Sui J / Lei J / Yang H / Brown AJ / Zhou Q / Yan C / Yan N
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 SREBP pathway controls cellular homeostasis of sterols. The key players in this pathway, Scap and Insig-1/2, are membrane-embedded sterol sensors. 25-hydroxycholesterol (25HC)-dependent ...The SREBP pathway controls cellular homeostasis of sterols. The key players in this pathway, Scap and Insig-1/2, are membrane-embedded sterol sensors. 25-hydroxycholesterol (25HC)-dependent association of Scap and Insigs acts as the master switch for the SREBP pathway. Here, we present cryo-EM 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 Å. The sterol sensing domain (SSD) in Scap and all six TMs in Insig-2 were resolved. A 25HC molecule is sandwiched between the S4-S6 segments in Scap and TMs 3/4 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.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionMar 6, 2020-
Header (metadata) releaseJan 20, 2021-
Map releaseJan 20, 2021-
UpdateJan 27, 2021-
Current statusJan 27, 2021Processing 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

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)
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
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

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Sample components

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Entire Scap and Insig complex

EntireName: Scap and Insig complex / Number of components: 4

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Component #1: protein, Scap and Insig complex

ProteinName: Scap and Insig complex / Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human) / Cell of expression system: HEK293F

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Component #2: protein, Insulin-induced gene 2 protein

ProteinName: Insulin-induced gene 2 protein / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 24.74966 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #3: protein, Sterol regulatory element-binding protein cleavage-activ...

ProteinName: Sterol regulatory element-binding protein cleavage-activating protein,Sterol regulatory element-binding protein cleavage-activating protein
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 63.295406 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Homo sapiens (human)

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Component #4: ligand, 25-HYDROXYCHOLESTEROL

LigandName: 25-HYDROXYCHOLESTEROL / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.402653 kDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 10 mg/mL / pH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 50 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: OTHER

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 153168
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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