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Yorodumi- PDB-6nzd: Cryo-EM Structure of the Lysosomal Folliculin Complex (FLCN-FNIP2... -
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-Basic information
Entry | Database: PDB / ID: 6nzd | ||||||
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Title | Cryo-EM Structure of the Lysosomal Folliculin Complex (FLCN-FNIP2-RagA-RagC-Ragulator) | ||||||
Components |
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Keywords | signaling protein/inhibitor / Lysosome / mTORC1 regulation / Amino acid sensing / GTPase / SIGNALING PROTEIN / signaling protein-inhibitor complex | ||||||
Function / homology | Function and homology information negative regulation of cell proliferation involved in kidney development / negative regulation of post-translational protein modification / cell proliferation involved in kidney development / regulation of cholesterol import / positive regulation of protein localization to lysosome / regulation of cell-substrate junction organization / Gtr1-Gtr2 GTPase complex / regulation of cholesterol efflux / positive regulation of RNA polymerase II regulatory region sequence-specific DNA binding / FNIP-folliculin RagC/D GAP ...negative regulation of cell proliferation involved in kidney development / negative regulation of post-translational protein modification / cell proliferation involved in kidney development / regulation of cholesterol import / positive regulation of protein localization to lysosome / regulation of cell-substrate junction organization / Gtr1-Gtr2 GTPase complex / regulation of cholesterol efflux / positive regulation of RNA polymerase II regulatory region sequence-specific DNA binding / FNIP-folliculin RagC/D GAP / Ragulator complex / negative regulation of brown fat cell differentiation / regulation of Ras protein signal transduction / protein localization to cell junction / regulation of TORC1 signaling / negative regulation of lysosome organization / regulation of pro-B cell differentiation / protein localization to lysosome / regulation of TOR signaling / TORC1 signaling / endosome organization / fibroblast migration / MTOR signalling / Amino acids regulate mTORC1 / lysosome localization / ATPase inhibitor activity / Energy dependent regulation of mTOR by LKB1-AMPK / kinase activator activity / protein localization to membrane / negative regulation of TOR signaling / enzyme-substrate adaptor activity / enzyme inhibitor activity / negative regulation of glycolytic process / cell-cell junction assembly / negative regulation of cold-induced thermogenesis / negative regulation of Rho protein signal transduction / endosomal transport / azurophil granule membrane / regulation of cell size / small GTPase-mediated signal transduction / Macroautophagy / lysosome organization / positive regulation of transforming growth factor beta receptor signaling pathway / RHOJ GTPase cycle / RHOQ GTPase cycle / mTORC1-mediated signalling / tertiary granule membrane / CDC42 GTPase cycle / hemopoiesis / RHOH GTPase cycle / ficolin-1-rich granule membrane / negative regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / centriolar satellite / RHOG GTPase cycle / TOR signaling / regulation of receptor recycling / positive regulation of TOR signaling / RAC2 GTPase cycle / RAC3 GTPase cycle / response to amino acid / cellular response to nutrient levels / positive regulation of autophagy / specific granule membrane / positive regulation of intrinsic apoptotic signaling pathway / protein-membrane adaptor activity / energy homeostasis / epithelial cell proliferation / Regulation of PTEN gene transcription / tumor necrosis factor-mediated signaling pathway / RAC1 GTPase cycle / positive regulation of TORC1 signaling / ERK1 and ERK2 cascade / phosphatidylinositol 3-kinase/protein kinase B signal transduction / cellular response to starvation / cellular response to amino acid starvation / intrinsic apoptotic signaling pathway / GTPase activator activity / viral genome replication / negative regulation of autophagy / cellular response to amino acid stimulus / transforming growth factor beta receptor signaling pathway / RNA splicing / negative regulation of cysteine-type endopeptidase activity involved in apoptotic process / guanyl-nucleotide exchange factor activity / cholesterol homeostasis / positive regulation of interleukin-8 production / regulation of cell growth / TP53 Regulates Metabolic Genes / phosphoprotein binding / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / regulation of protein phosphorylation / MAP2K and MAPK activation / positive regulation of protein-containing complex assembly / response to virus / cilium / negative regulation of ERK1 and ERK2 cascade / mitotic spindle / positive regulation of protein localization to nucleus / GDP binding / negative regulation of epithelial cell proliferation Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | ||||||
Authors | Fromm, S.A. / Young, L.N. / Hurley, J.H. | ||||||
Funding support | United States, 1items
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Citation | Journal: Science / Year: 2019 Title: Structural mechanism of a Rag GTPase activation checkpoint by the lysosomal folliculin complex. Authors: Rosalie E Lawrence / Simon A Fromm / Yangxue Fu / Adam L Yokom / Do Jin Kim / Ashley M Thelen / Lindsey N Young / Chun-Yan Lim / Avi J Samelson / James H Hurley / Roberto Zoncu / Abstract: The tumor suppressor folliculin (FLCN) enables nutrient-dependent activation of the mechanistic target of rapamycin complex 1 (mTORC1) protein kinase via its guanosine triphosphatase (GTPase) ...The tumor suppressor folliculin (FLCN) enables nutrient-dependent activation of the mechanistic target of rapamycin complex 1 (mTORC1) protein kinase via its guanosine triphosphatase (GTPase) activating protein (GAP) activity toward the GTPase RagC. Concomitant with mTORC1 inactivation by starvation, FLCN relocalizes from the cytosol to lysosomes. To determine the lysosomal function of FLCN, we reconstituted the human lysosomal FLCN complex (LFC) containing FLCN, its partner FLCN-interacting protein 2 (FNIP2), and the RagA:RagC GTPases as they exist in the starved state with their lysosomal anchor Ragulator complex and determined its cryo-electron microscopy structure to 3.6 angstroms. The RagC-GAP activity of FLCN was inhibited within the LFC, owing to displacement of a catalytically required arginine in FLCN from the RagC nucleotide. Disassembly of the LFC and release of the RagC-GAP activity of FLCN enabled mTORC1-dependent regulation of the master regulator of lysosomal biogenesis, transcription factor E3, implicating the LFC as a checkpoint in mTORC1 signaling. | ||||||
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 | 6nzd.cif.gz | 350.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6nzd.ent.gz | 265.2 KB | Display | PDB format |
PDBx/mmJSON format | 6nzd.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6nzd_validation.pdf.gz | 901.4 KB | Display | wwPDB validaton report |
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Full document | 6nzd_full_validation.pdf.gz | 925.9 KB | Display | |
Data in XML | 6nzd_validation.xml.gz | 60.8 KB | Display | |
Data in CIF | 6nzd_validation.cif.gz | 91.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/nz/6nzd ftp://data.pdbj.org/pub/pdb/validation_reports/nz/6nzd | HTTPS FTP |
-Related structure data
Related structure data | 0554MC 0556C C: citing same article (ref.) M: map data used to model this data |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Ragulator complex protein ... , 4 types, 4 molecules ABCD
#1: Protein | Mass: 18325.350 Da / Num. of mol.: 1 / Mutation: G2A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR1, C11orf59, PDRO, PP7157 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q6IAA8 |
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#2: Protein | Mass: 13645.579 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR2, MAPBPIP, ROBLD3, HSPC003 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9Y2Q5 |
#3: Protein | Mass: 13637.678 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR3, MAP2K1IP1, MAPKSP1, PRO2783 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9UHA4 |
#4: Protein | Mass: 10753.236 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR4, C7orf59 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q0VGL1 |
-Protein , 3 types, 3 molecules EHI
#5: Protein | Mass: 18178.520 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: LAMTOR5, HBXIP, hCG_40252 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: A0A0C4DGV4, UniProt: O43504*PLUS |
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#8: Protein | Mass: 69143.070 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: FLCN, BHD / Cell line (production host): HEK 293 GNTI / Production host: Homo sapiens (human) / References: UniProt: Q8NFG4 |
#9: Protein | Mass: 122475.414 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: FNIP2, FNIPL, KIAA1450, MAPO1 / Cell line (production host): HEK 293 GNTI / Production host: Homo sapiens (human) / References: UniProt: Q9P278 |
-Ras-related GTP-binding protein ... , 2 types, 2 molecules FG
#6: Protein | Mass: 36615.168 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RRAGA / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q7L523 |
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#7: Protein | Mass: 44758.336 Da / Num. of mol.: 1 / Mutation: D181N Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RRAGC / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9HB90 |
-Non-polymers , 2 types, 2 molecules
#10: Chemical | ChemComp-GDP / |
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#11: Chemical | ChemComp-L8S / |
-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: FLCN-FNIP2-RagA-RagC-Ragulator Complex / Type: COMPLEX / Details: RagA bound to GDP; RagC bound to XTPgammaS / Entity ID: #1-#9 / Source: MULTIPLE SOURCES | ||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.34 MDa / Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) | Organism: Homo sapiens (human) | ||||||||||||||||||||||||||||||
Source (recombinant) | Organism: Spodoptera frugiperda (fall armyworm) | ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.4 | ||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||
Specimen support | Details: 15 W / Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: C-flat-2/1 | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 90 % / Chamber temperature: 277 K / Details: Whatman 597 |
-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: OTHER |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 11 sec. / Electron dose: 65.6 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2703 |
EM imaging optics | Energyfilter slit width: 20 eV |
Image scans | Movie frames/image: 44 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 982343 | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 163376 / Symmetry type: POINT | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building | Space: REAL | |||||||||||||||||||||||||||||||||||||||||||||||||||||||
Atomic model building |
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Refinement | Highest resolution: 3.6 Å |