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- EMDB-3927: Architecture of the human mTORC2 core complex (C2) -

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Basic information

Entry
Database: EMDB / ID: EMD-3927
TitleArchitecture of the human mTORC2 core complex (C2)
Map dataHuman mTORC2 core complex (C2)
Sample
  • Complex: Human mTORC2 Core Complex (C2)
Function / homology
Function and homology information


: / TORC2 signaling / : / regulation of peptidyl-serine phosphorylation / RNA polymerase III type 2 promoter sequence-specific DNA binding / positive regulation of cytoplasmic translational initiation / RNA polymerase III type 1 promoter sequence-specific DNA binding / positive regulation of pentose-phosphate shunt / T-helper 1 cell lineage commitment / regulation of locomotor rhythm ...: / TORC2 signaling / : / regulation of peptidyl-serine phosphorylation / RNA polymerase III type 2 promoter sequence-specific DNA binding / positive regulation of cytoplasmic translational initiation / RNA polymerase III type 1 promoter sequence-specific DNA binding / positive regulation of pentose-phosphate shunt / T-helper 1 cell lineage commitment / regulation of locomotor rhythm / positive regulation of wound healing, spreading of epidermal cells / cellular response to leucine starvation / TFIIIC-class transcription factor complex binding / TORC2 complex / heart valve morphogenesis / regulation of membrane permeability / negative regulation of lysosome organization / RNA polymerase III type 3 promoter sequence-specific DNA binding / TORC1 complex / positive regulation of transcription of nucleolar large rRNA by RNA polymerase I / calcineurin-NFAT signaling cascade / regulation of cellular response to oxidative stress / regulation of autophagosome assembly / TORC1 signaling / voluntary musculoskeletal movement / regulation of osteoclast differentiation / positive regulation of keratinocyte migration / cellular response to L-leucine / MTOR signalling / Amino acids regulate mTORC1 / cellular response to nutrient / energy reserve metabolic process / Energy dependent regulation of mTOR by LKB1-AMPK / phosphatidic acid binding / nucleus localization / negative regulation of Ras protein signal transduction / ruffle organization / negative regulation of cell size / cellular response to osmotic stress / phosphatidylinositol-3,4-bisphosphate binding / phosphatidylinositol-3,5-bisphosphate binding / regulation of establishment of cell polarity / anoikis / cardiac muscle cell development / positive regulation of transcription by RNA polymerase III / negative regulation of protein localization to nucleus / embryo development ending in birth or egg hatching / regulation of myelination / negative regulation of calcineurin-NFAT signaling cascade / Macroautophagy / regulation of cell size / negative regulation of macroautophagy / lysosome organization / positive regulation of oligodendrocyte differentiation / positive regulation of actin filament polymerization / positive regulation of myotube differentiation / behavioral response to pain / regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / TOR signaling / oligodendrocyte differentiation / mTORC1-mediated signalling / germ cell development / Constitutive Signaling by AKT1 E17K in Cancer / phosphatidylinositol-3,4,5-trisphosphate binding / cellular response to nutrient levels / CD28 dependent PI3K/Akt signaling / positive regulation of phosphoprotein phosphatase activity / positive regulation of translational initiation / neuronal action potential / HSF1-dependent transactivation / positive regulation of TOR signaling / positive regulation of epithelial to mesenchymal transition / regulation of macroautophagy / endomembrane system / 'de novo' pyrimidine nucleobase biosynthetic process / response to amino acid / positive regulation of lipid biosynthetic process / phagocytic vesicle / positive regulation of lamellipodium assembly / heart morphogenesis / regulation of cellular response to heat / cytoskeleton organization / cardiac muscle contraction / positive regulation of stress fiber assembly / phosphatidylinositol-4,5-bisphosphate binding / cellular response to amino acid starvation / T cell costimulation / substantia nigra development / cellular response to starvation / positive regulation of endothelial cell proliferation / positive regulation of glycolytic process / protein serine/threonine kinase activator activity / response to nutrient levels / post-embryonic development / negative regulation of autophagy / response to nutrient / VEGFR2 mediated vascular permeability / positive regulation of translation / regulation of signal transduction by p53 class mediator / Regulation of PTEN gene transcription
Similarity search - Function
TORC2 component Bit61/PRR5 / HbrB-like / Rapamycin-insensitive companion of mTOR, N-terminal domain / Pianissimo family / Rapamycin-insensitive companion of mTOR, phosphorylation-site / Rapamycin-insensitive companion of mTOR, middle domain / Rapamycin-insensitive companion of mTOR, domain 5 / Rapamycin-insensitive companion of mTOR, domain 4 / Rapamycin-insensitive companion of mTOR RasGEF_N domain / Rapamycin-insensitive companion of mTOR, N-term ...TORC2 component Bit61/PRR5 / HbrB-like / Rapamycin-insensitive companion of mTOR, N-terminal domain / Pianissimo family / Rapamycin-insensitive companion of mTOR, phosphorylation-site / Rapamycin-insensitive companion of mTOR, middle domain / Rapamycin-insensitive companion of mTOR, domain 5 / Rapamycin-insensitive companion of mTOR, domain 4 / Rapamycin-insensitive companion of mTOR RasGEF_N domain / Rapamycin-insensitive companion of mTOR, N-term / Rapamycin-insensitive companion of mTOR, phosphorylation-site / Rapamycin-insensitive companion of mTOR, middle domain / Rapamycin-insensitive companion of mTOR, domain 5 / Rapamycin-insensitive companion of mTOR RasGEF_N domain / Rapamycin-insensitive companion of mTOR, middle domain / Rapamycin-insensitive companion of mTOR, N-term / Rapamycin-insensitive companion of mTOR, phosphorylation-site / Rapamycin-insensitive companion of mTOR, domain 5 / Sin1, N-terminal / Stress-activated map kinase interacting protein 1 (SIN1) / TORC2 component Sin1/Avo1 / SAPK-interacting protein 1, Pleckstrin-homology domain / Sin1, middle CRIM domain / SAPK-interacting protein 1 (Sin1), middle CRIM domain / SAPK-interacting protein 1 (Sin1), Pleckstrin-homology / Target of rapamycin complex subunit LST8 / Domain of unknown function DUF3385, target of rapamycin protein / Domain of unknown function (DUF3385) / 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 / Cullin repeat-like-containing domain superfamily / PIK-related kinase, FAT / FAT domain / FATC 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 / PH-like 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 / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40/YVTN repeat-like-containing domain superfamily / Protein kinase-like domain superfamily
Similarity search - Domain/homology
Serine/threonine-protein kinase mTOR / Proline-rich protein 5 / Rapamycin-insensitive companion of mTOR / Target of rapamycin complex 2 subunit MAPKAP1 / Target of rapamycin complex subunit LST8
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.4 Å
AuthorsAylett CHS / Boehringer D / Stuttfeld E / Imseng S / Scaiola A / Sauer E / Hall MN / Maier T / Ban N
CitationJournal: Elife / Year: 2018
Title: Architecture of the human mTORC2 core complex.
Authors: Edward Stuttfeld / Christopher Hs Aylett / Stefan Imseng / Daniel Boehringer / Alain Scaiola / Evelyn Sauer / Michael N Hall / Timm Maier / Nenad Ban /
Abstract: The mammalian target of rapamycin (mTOR) is a key protein kinase controlling cellular metabolism and growth. It is part of the two structurally and functionally distinct multiprotein complexes mTORC1 ...The mammalian target of rapamycin (mTOR) is a key protein kinase controlling cellular metabolism and growth. It is part of the two structurally and functionally distinct multiprotein complexes mTORC1 and mTORC2. Dysregulation of mTOR occurs in diabetes, cancer and neurological disease. We report the architecture of human mTORC2 at intermediate resolution, revealing a conserved binding site for accessory proteins on mTOR and explaining the structural basis for the rapamycin insensitivity of the complex.
History
DepositionOct 15, 2017-
Header (metadata) releaseDec 20, 2017-
Map releaseFeb 28, 2018-
UpdateFeb 28, 2018-
Current statusFeb 28, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_3927.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationHuman mTORC2 core complex (C2)
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.04 / Movie #1: 0.04
Minimum - Maximum-0.021306371 - 0.094786234
Average (Standard dev.)0.0007714506 (±0.0057059457)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 407.03998 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z407.040407.040407.040
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-0.0210.0950.001

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Supplemental data

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Sample components

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Entire : Human mTORC2 Core Complex (C2)

EntireName: Human mTORC2 Core Complex (C2)
Components
  • Complex: Human mTORC2 Core Complex (C2)

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Supramolecule #1: Human mTORC2 Core Complex (C2)

SupramoleculeName: Human mTORC2 Core Complex (C2) / type: complex / ID: 1 / Parent: 0
Details: Core complex of mTORC2 comprising; mTOR, Rictor, mLST8, mSIN1 and Protor-1
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm) / Recombinant cell: Sf9 / Recombinant plasmid: Multibac
Molecular weightTheoretical: 1 MDa

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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

BufferpH: 8 / Details: 150 mM NaCl, 15 mM NaBicine pH 8.0, 1 mM TCEP
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 95 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: Two seconds blotting.
DetailsSample was prepared by a modified GRAFIX protocol described in the paper, and was extremely dilute. Particles were adhered to a thin carbon film to obtain reasonable coverage.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Calibrated magnification: 47100 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.0 µm
Specialist opticsEnergy filter - Name: GIF Quantum SE
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
TemperatureMin: 100.0 K / Max: 100.0 K
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Digitization - Sampling interval: 5.0 µm / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Number real images: 3997 / Average exposure time: 20.0 sec. / Average electron dose: 80.0 e/Å2 / Details: Dose weighting applied
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 207920
CTF correctionSoftware - Name: CTFFIND (ver. 4) / Details: Performed in Relion
Startup modelType of model: OTHER
Details: Generated from negatively stained sample using EMAN2
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.0) / Details: RELION initial cross-correlation search
Final 3D classificationNumber classes: 6 / Software - Name: RELION (ver. 2.0) / Details: Relion MAP
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.0)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C2 (2 fold cyclic) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 7.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.0) / Number images used: 17183
FSC plot (resolution estimation)

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