+Open data
-Basic information
Entry | Database: PDB / ID: 6uox | |||||||||
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Title | Structure of itraconazole-bound NPC1 | |||||||||
Components | NPC intracellular cholesterol transporter 1 | |||||||||
Keywords | MEMBRANE PROTEIN / Niemann-Pick C disease / cholesterol transport / sterol-sensing domain | |||||||||
Function / homology | Function and homology information cyclodextrin metabolic process / cholesterol storage / membrane raft organization / : / intracellular cholesterol transport / intracellular lipid transport / intestinal cholesterol absorption / LDL clearance / negative regulation of epithelial cell apoptotic process / bile acid metabolic process ...cyclodextrin metabolic process / cholesterol storage / membrane raft organization / : / intracellular cholesterol transport / intracellular lipid transport / intestinal cholesterol absorption / LDL clearance / negative regulation of epithelial cell apoptotic process / bile acid metabolic process / cholesterol transport / programmed cell death / establishment of protein localization to membrane / adult walking behavior / lysosomal transport / cholesterol efflux / cellular response to steroid hormone stimulus / negative regulation of macroautophagy / cholesterol binding / protein glycosylation / cellular response to low-density lipoprotein particle stimulus / response to cadmium ion / negative regulation of TORC1 signaling / cholesterol metabolic process / neurogenesis / liver development / cholesterol homeostasis / macroautophagy / autophagy / endocytosis / transmembrane signaling receptor activity / late endosome membrane / nuclear envelope / virus receptor activity / signaling receptor activity / gene expression / lysosome / response to xenobiotic stimulus / symbiont entry into host cell / membrane raft / lysosomal membrane / perinuclear region of cytoplasm / Golgi apparatus / endoplasmic reticulum / extracellular exosome / extracellular region / membrane / plasma membrane Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.02 Å | |||||||||
Authors | Long, T. / Li, X. | |||||||||
Citation | Journal: Nat Commun / Year: 2020 Title: Structural basis for itraconazole-mediated NPC1 inhibition. Authors: Tao Long / Xiaofeng Qi / Abdirahman Hassan / Qiren Liang / Jef K De Brabander / Xiaochun Li / Abstract: Niemann-Pick C1 (NPC1), a lysosomal protein of 13 transmembrane helices (TMs) and three lumenal domains, exports low-density-lipoprotein (LDL)-derived cholesterol from lysosomes. TMs 3-7 of NPC1 ...Niemann-Pick C1 (NPC1), a lysosomal protein of 13 transmembrane helices (TMs) and three lumenal domains, exports low-density-lipoprotein (LDL)-derived cholesterol from lysosomes. TMs 3-7 of NPC1 comprise the Sterol-Sensing Domain (SSD). Previous studies suggest that mutation of the NPC1-SSD or the addition of the anti-fungal drug itraconazole abolishes NPC1 activity in cells. However, the itraconazole binding site and the mechanism of NPC1-mediated cholesterol transport remain unknown. Here, we report a cryo-EM structure of human NPC1 bound to itraconazole, which reveals how this binding site in the center of NPC1 blocks a putative lumenal tunnel linked to the SSD. Functional assays confirm that blocking this tunnel abolishes NPC1-mediated cholesterol egress. Intriguingly, the palmitate anchor of Hedgehog occupies a similar site in the homologous tunnel of Patched, suggesting a conserved mechanism for sterol transport in this family of proteins and establishing a central function of their SSDs. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6uox.cif.gz | 240.2 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6uox.ent.gz | 190.3 KB | Display | PDB format |
PDBx/mmJSON format | 6uox.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6uox_validation.pdf.gz | 984.1 KB | Display | wwPDB validaton report |
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Full document | 6uox_full_validation.pdf.gz | 989.6 KB | Display | |
Data in XML | 6uox_validation.xml.gz | 36.2 KB | Display | |
Data in CIF | 6uox_validation.cif.gz | 55.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/uo/6uox ftp://data.pdbj.org/pub/pdb/validation_reports/uo/6uox | HTTPS FTP |
-Related structure data
Related structure data | 20834MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 143315.016 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: NPC1 / Production host: Homo sapiens (human) / References: UniProt: O15118 | ||||
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#2: Polysaccharide | 2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source | ||||
#3: Polysaccharide | alpha-D-mannopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta- ...alpha-D-mannopyranose-(1-3)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source | ||||
#4: Sugar | ChemComp-NAG / #5: Chemical | ChemComp-QDG / | Has ligand of interest | Y | |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: NPC1 and itraconazole-Br complex / Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT |
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Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Homo sapiens (human) |
Buffer solution | pH: 7.4 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: DARK FIELD |
Image recording | Electron dose: 80 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
Software | Name: REFMAC / Version: 5.8.0135 / Classification: refinement | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 4.02 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 209612 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refinement | Resolution: 4.13→231 Å / Cor.coef. Fo:Fc: 0.832 / SU B: 59.618 / SU ML: 0.778 / ESU R: 0.305 / Stereochemistry target values: MAXIMUM LIKELIHOOD / Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
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Solvent computation | Ion probe radii: 0.8 Å / Shrinkage radii: 0.8 Å / VDW probe radii: 1.2 Å / Solvent model: MASK | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 175.814 Å2
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Refinement step | Cycle: 1 / Total: 9138 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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