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- PDB-5h64: Cryo-EM structure of mTORC1 -

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Basic information

Entry
Database: PDB / ID: 5h64
TitleCryo-EM structure of mTORC1
Components
  • Regulatory-associated protein of mTOR
  • Serine/threonine-protein kinase mTOR
  • Target of rapamycin complex subunit LST8
KeywordsTRANSFERASE / cryo structure mTOR complex
Function / homology
Function and homology information


RNA polymerase III type 2 promoter sequence-specific DNA binding / RNA polymerase III type 1 promoter sequence-specific DNA binding / positive regulation of cytoplasmic translational initiation / T-helper 1 cell lineage commitment / positive regulation of pentose-phosphate shunt / regulation of locomotor rhythm / positive regulation of wound healing, spreading of epidermal cells / TORC2 signaling / TORC2 complex / regulation of membrane permeability ...RNA polymerase III type 2 promoter sequence-specific DNA binding / RNA polymerase III type 1 promoter sequence-specific DNA binding / positive regulation of cytoplasmic translational initiation / T-helper 1 cell lineage commitment / positive regulation of pentose-phosphate shunt / regulation of locomotor rhythm / positive regulation of wound healing, spreading of epidermal cells / TORC2 signaling / TORC2 complex / regulation of membrane permeability / cellular response to leucine starvation / heart valve morphogenesis / negative regulation of lysosome organization / TFIIIC-class transcription factor complex binding / TORC1 complex / positive regulation of transcription of nucleolar large rRNA by RNA polymerase I / calcineurin-NFAT signaling cascade / voluntary musculoskeletal movement / positive regulation of odontoblast differentiation / regulation of osteoclast differentiation / RNA polymerase III type 3 promoter sequence-specific DNA binding / positive regulation of keratinocyte migration / regulation of lysosome organization / Amino acids regulate mTORC1 / cellular response to L-leucine / MTOR signalling / cellular response to nutrient / regulation of autophagosome assembly / Energy dependent regulation of mTOR by LKB1-AMPK / TORC1 signaling / energy reserve metabolic process / ruffle organization / serine/threonine protein kinase complex / negative regulation of cell size / cellular response to methionine / positive regulation of osteoclast differentiation / positive regulation of ubiquitin-dependent protein catabolic process / inositol hexakisphosphate binding / cellular response to osmotic stress / anoikis / protein serine/threonine kinase inhibitor activity / negative regulation of protein localization to nucleus / cardiac muscle cell development / negative regulation of calcineurin-NFAT signaling cascade / regulation of myelination / positive regulation of transcription by RNA polymerase III / positive regulation of peptidyl-threonine phosphorylation / positive regulation of actin filament polymerization / negative regulation of macroautophagy / Macroautophagy / regulation of cell size / positive regulation of myotube differentiation / Constitutive Signaling by AKT1 E17K in Cancer / social behavior / oligodendrocyte differentiation / germ cell development / behavioral response to pain / TOR signaling / mTORC1-mediated signalling / positive regulation of oligodendrocyte differentiation / positive regulation of translational initiation / protein kinase activator activity / CD28 dependent PI3K/Akt signaling / positive regulation of TOR signaling / response to amino acid / HSF1-dependent transactivation / positive regulation of G1/S transition of mitotic cell cycle / regulation of macroautophagy / enzyme-substrate adaptor activity / 'de novo' pyrimidine nucleobase biosynthetic process / cellular response to nutrient levels / vascular endothelial cell response to laminar fluid shear stress / neuronal action potential / positive regulation of lipid biosynthetic process / positive regulation of epithelial to mesenchymal transition / heart morphogenesis / regulation of cellular response to heat / positive regulation of lamellipodium assembly / cardiac muscle contraction / positive regulation of peptidyl-serine phosphorylation / phagocytic vesicle / positive regulation of stress fiber assembly / positive regulation of endothelial cell proliferation / 14-3-3 protein binding / T cell costimulation / cytoskeleton organization / endomembrane system / negative regulation of autophagy / cellular response to amino acid starvation / protein serine/threonine kinase activator activity / positive regulation of glycolytic process / cellular response to starvation / regulation of signal transduction by p53 class mediator / Regulation of PTEN gene transcription / post-embryonic development / positive regulation of translation / VEGFR2 mediated vascular permeability / TP53 Regulates Metabolic Genes / regulation of actin cytoskeleton organization / phosphoprotein binding
Similarity search - Function
Raptor, N-terminal CASPase-like domain / Raptor N-terminal CASPase like domain / Raptor N-terminal CASPase like domain / Regulatory associated protein of TOR / Target of rapamycin complex subunit LST8 / Domain of unknown function DUF3385, target of rapamycin protein / Serine/threonine-protein kinase mTOR domain / Domain of unknown function / FKBP12-rapamycin binding domain / Serine/threonine-protein kinase TOR ...Raptor, N-terminal CASPase-like domain / Raptor N-terminal CASPase like domain / Raptor N-terminal CASPase like domain / Regulatory associated protein of TOR / Target of rapamycin complex subunit LST8 / Domain of unknown function DUF3385, target of rapamycin protein / Serine/threonine-protein kinase mTOR domain / Domain of unknown function / FKBP12-rapamycin binding domain / Serine/threonine-protein kinase TOR / FKBP12-rapamycin binding domain superfamily / FKBP12-rapamycin binding domain / Rapamycin binding domain / Serine/threonine-protein kinase ATR-like, HEAT repeats / HEAT repeat / HEAT repeat / : / FATC domain / PIK-related kinase, FAT / FAT domain / FATC / FATC domain / PIK-related kinase / FAT domain profile. / FATC domain profile. / Quinoprotein alcohol dehydrogenase-like superfamily / Phosphatidylinositol 3- and 4-kinases signature 1. / Phosphatidylinositol 3/4-kinase, conserved site / Phosphatidylinositol 3- and 4-kinases signature 2. / Phosphatidylinositol 3-/4-kinase, catalytic domain superfamily / Phosphoinositide 3-kinase, catalytic domain / Phosphatidylinositol 3- and 4-kinase / Phosphatidylinositol 3- and 4-kinases catalytic domain profile. / Phosphatidylinositol 3-/4-kinase, catalytic domain / Armadillo-like helical / Tetratricopeptide-like helical domain superfamily / Armadillo-type fold / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / WD domain, G-beta repeat / 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 / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Serine/threonine-protein kinase mTOR / Regulatory-associated protein of mTOR / Target of rapamycin complex subunit LST8
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.4 Å
AuthorsYang, H. / Wang, J. / Liu, M. / Chen, X. / Huang, M. / Tan, D. / Dong, M. / Wong, C.C.L. / Wang, J. / Xu, Y. / Wang, H.
Funding support China, 5items
OrganizationGrant numberCountry
National Natural Science Foundation of ChinaU1432242 China
National Natural Science Foundation of China31425008 China
National Natural Science Foundation of China91419301 China
Basic Research Project of Shanghai Science and Technology Commission12JC1402700 China
Program of Shanghai Subject Chief Scientist14XD1400500 China
CitationJournal: Protein Cell / Year: 2016
Title: 4.4 Å Resolution Cryo-EM structure of human mTOR Complex 1.
Authors: Huirong Yang / Jia Wang / Mengjie Liu / Xizi Chen / Min Huang / Dan Tan / Meng-Qiu Dong / Catherine C L Wong / Jiawei Wang / Yanhui Xu / Hong-Wei Wang /
Abstract: Mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) integrates signals from growth factors, cellular energy levels, stress and amino acids to control cell growth and proliferation through ...Mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) integrates signals from growth factors, cellular energy levels, stress and amino acids to control cell growth and proliferation through regulating translation, autophagy and metabolism. Here we determined the cryo-electron microscopy structure of human mTORC1 at 4.4 Å resolution. The mTORC1 comprises a dimer of heterotrimer (mTOR-Raptor-mLST8) mediated by the mTOR protein. The complex adopts a hollow rhomboid shape with 2-fold symmetry. Notably, mTORC1 shows intrinsic conformational dynamics. Within the complex, the conserved N-terminal caspase-like domain of Raptor faces toward the catalytic cavity of the kinase domain of mTOR. Raptor shows no caspase activity and therefore may bind to TOS motif for substrate recognition. Structural analysis indicates that FKBP12-Rapamycin may generate steric hindrance for substrate entry to the catalytic cavity of mTORC1. The structure provides a basis to understand the assembly of mTORC1 and a framework to characterize the regulatory mechanism of mTORC1 pathway.
History
DepositionNov 10, 2016Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 25, 2017Provider: repository / Type: Initial release
Revision 1.1Mar 1, 2017Group: Database references
Revision 1.2Dec 6, 2017Group: Advisory / Data processing / Database references
Category: citation / citation_author ...citation / citation_author / em_software / pdbx_unobs_or_zero_occ_atoms
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _em_software.name
Revision 1.3Nov 6, 2019Group: Data collection / Other / Category: atom_sites / cell
Item: _atom_sites.fract_transf_matrix[1][1] / _atom_sites.fract_transf_matrix[2][2] ..._atom_sites.fract_transf_matrix[1][1] / _atom_sites.fract_transf_matrix[2][2] / _atom_sites.fract_transf_matrix[3][3] / _cell.Z_PDB
Revision 1.4Oct 9, 2024Group: Data collection / Database references / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_admin.last_update

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Structure visualization

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  • Deposited structure unit
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Structure viewerMolecule:
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Assembly

Deposited unit
A: Serine/threonine-protein kinase mTOR
B: Regulatory-associated protein of mTOR
C: Target of rapamycin complex subunit LST8
a: Serine/threonine-protein kinase mTOR
b: Regulatory-associated protein of mTOR
c: Target of rapamycin complex subunit LST8


Theoretical massNumber of molelcules
Total (without water)948,7366
Polymers948,7366
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area10360 Å2
ΔGint-60 kcal/mol
Surface area299360 Å2

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Components

#1: Protein Serine/threonine-protein kinase mTOR / FK506-binding protein 12-rapamycin complex-associated protein 1 / FKBP12-rapamycin complex- ...FK506-binding protein 12-rapamycin complex-associated protein 1 / FKBP12-rapamycin complex-associated protein / Mammalian target of rapamycin / mTOR / Mechanistic target of rapamycin / Rapamycin and FKBP12 target 1 / Rapamycin target protein 1


Mass: 289257.969 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MTOR, FRAP, FRAP1, FRAP2, RAFT1, RAPT1 / Cell line (production host): HEK293F / Production host: Homo sapiens (human)
References: UniProt: P42345, non-specific serine/threonine protein kinase
#2: Protein Regulatory-associated protein of mTOR / Raptor / p150 target of rapamycin (TOR)-scaffold protein


Mass: 149200.016 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: RPTOR, KIAA1303, RAPTOR / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q8N122
#3: Protein Target of rapamycin complex subunit LST8 / TORC subunit LST8 / G protein beta subunit-like / Protein GbetaL / Mammalian lethal with SEC13 ...TORC subunit LST8 / G protein beta subunit-like / Protein GbetaL / Mammalian lethal with SEC13 protein 8 / mLST8


Mass: 35910.090 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: MLST8, GBL, LST8 / Cell line (production host): HEK293F / Production host: Homo sapiens (human) / References: UniProt: Q9BVC4
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: human mTOR complex 1 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 1 MDa / Experimental value: YES
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Mammalia (mammals) / Plasmid: pCAG
Buffer solutionpH: 7.4
Buffer component
IDConc.NameBuffer-ID
120 mMHEPES1
2150 mMNaCl1
31 mMTCEP1
SpecimenConc.: 1.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil
VitrificationCryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 295 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELD / Nominal magnification: 22500 X / Calibrated magnification: 22500 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Calibrated defocus min: 1500 nm / Calibrated defocus max: 2500 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 0.25 sec. / Electron dose: 8.25 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 2997
Image scansWidth: 7676 / Height: 7420 / Movie frames/image: 32 / Used frames/image: 0-31

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Processing

SoftwareName: PHENIX / Version: 1.10.1_2155: / Classification: refinement
EM software
IDNameVersionCategory
1RELION1.4particle selection
2UCSFImage4image acquisition
4CTFFIND33CTF correction
7EMfitmodel fitting
9RELION1.4initial Euler assignment
10RELION1.4final Euler assignment
12RELION1.43D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 486584
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 115039 / Algorithm: BACK PROJECTION / Symmetry type: POINT
Atomic model buildingProtocol: OTHER
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.01149020
ELECTRON MICROSCOPYf_angle_d1.46666566
ELECTRON MICROSCOPYf_dihedral_angle_d8.46839286
ELECTRON MICROSCOPYf_chiral_restr0.0647738
ELECTRON MICROSCOPYf_plane_restr0.0088636

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