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Yorodumi- PDB-7nzm: Cryo-EM structure of pre-dephosphorylation complex of phosphoryla... -
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-Basic information
Entry | Database: PDB / ID: 7nzm | ||||||
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Title | Cryo-EM structure of pre-dephosphorylation complex of phosphorylated eIF2alpha with trapped holophosphatase (PP1A_D64A/PPP1R15A/G-actin/DNase I) | ||||||
Components |
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Keywords | HYDROLASE / holophosphatase / PP1 / PPP1R15A / phosphorylated eIF2alpha | ||||||
Function / homology | Function and homology information fatty acid derivative binding / positive regulation of translational initiation in response to stress / : / regulation of translational initiation by eIF2 alpha dephosphorylation / regulation of neutrophil mediated cytotoxicity / regulation of acute inflammatory response / zymogen granule / : / protein phosphatase type 1 complex / : ...fatty acid derivative binding / positive regulation of translational initiation in response to stress / : / regulation of translational initiation by eIF2 alpha dephosphorylation / regulation of neutrophil mediated cytotoxicity / regulation of acute inflammatory response / zymogen granule / : / protein phosphatase type 1 complex / : / regulation of translation in response to endoplasmic reticulum stress / translation initiation ternary complex / glial limiting end-foot / deoxyribonuclease I / HRI-mediated signaling / response to kainic acid / Cellular response to mitochondrial stress / response to manganese-induced endoplasmic reticulum stress / PTW/PP1 phosphatase complex / positive regulation of type B pancreatic cell apoptotic process / negative regulation of translational initiation in response to stress / Response of EIF2AK1 (HRI) to heme deficiency / Recycling of eIF2:GDP / PERK-mediated unfolded protein response / peptidyl-serine dephosphorylation / PERK regulates gene expression / regulation of translational initiation in response to stress / eukaryotic translation initiation factor 2 complex / : / protein localization to endoplasmic reticulum / deoxyribonuclease I activity / neutrophil activation involved in immune response / protein phosphatase 1 binding / protein phosphatase regulator activity / eukaryotic 48S preinitiation complex / DNA catabolic process / Formation of the ternary complex, and subsequently, the 43S complex / cytoskeletal motor activator activity / Ribosomal scanning and start codon recognition / glycogen metabolic process / protein serine/threonine phosphatase activity / Translation initiation complex formation / myosin phosphatase activity / tropomyosin binding / protein-serine/threonine phosphatase / myosin heavy chain binding / negative regulation of PERK-mediated unfolded protein response / mesenchyme migration / detection of maltose stimulus / entrainment of circadian clock by photoperiod / troponin I binding / maltose transport complex / protein phosphatase activator activity / filamentous actin / actin filament bundle / phosphatase activity / carbohydrate transport / : / skeletal muscle thin filament assembly / actin filament bundle assembly / phosphoprotein phosphatase activity / striated muscle thin filament / maltose binding / transition metal ion binding / Response of EIF2AK4 (GCN2) to amino acid deficiency / maltose transport / intrinsic apoptotic signaling pathway in response to endoplasmic reticulum stress / skeletal muscle myofibril / maltodextrin transmembrane transport / actin monomer binding / carbohydrate transmembrane transporter activity / mitophagy / GTP hydrolysis and joining of the 60S ribosomal subunit / L13a-mediated translational silencing of Ceruloplasmin expression / ATP-binding cassette (ABC) transporter complex, substrate-binding subunit-containing / skeletal muscle fiber development / stress fiber / stress granule assembly / titin binding / response to endoplasmic reticulum stress / protein dephosphorylation / translation initiation factor activity / cellular response to amino acid starvation / actin filament polymerization / ATP-binding cassette (ABC) transporter complex / cell chemotaxis / Downregulation of TGF-beta receptor signaling / filopodium / actin filament / translational initiation / circadian regulation of gene expression / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / regulation of circadian rhythm / ABC-family proteins mediated transport / PKR-mediated signaling / cytoplasmic stress granule / calcium-dependent protein binding / cellular response to UV / positive regulation of canonical Wnt signaling pathway / lamellipodium Similarity search - Function | ||||||
Biological species | Homo sapiens (human) Oryctolagus cuniculus (rabbit) Escherichia coli (E. coli) Bos taurus (cattle) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.96 Å | ||||||
Authors | Yan, Y. / Hardwick, S. / Ron, D. | ||||||
Funding support | United Kingdom, 1items
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Citation | Journal: Nat Struct Mol Biol / Year: 2021 Title: Higher-order phosphatase-substrate contacts terminate the integrated stress response. Authors: Yahui Yan / Heather P Harding / David Ron / Abstract: Many regulatory PPP1R subunits join few catalytic PP1c subunits to mediate phosphoserine and phosphothreonine dephosphorylation in metazoans. Regulatory subunits engage the surface of PP1c, locally ...Many regulatory PPP1R subunits join few catalytic PP1c subunits to mediate phosphoserine and phosphothreonine dephosphorylation in metazoans. Regulatory subunits engage the surface of PP1c, locally affecting flexible access of the phosphopeptide to the active site. However, catalytic efficiency of holophosphatases towards their phosphoprotein substrates remains unexplained. Here we present a cryo-EM structure of the tripartite PP1c-PPP1R15A-G-actin holophosphatase that terminates signaling in the mammalian integrated stress response (ISR) in the pre-dephosphorylation complex with its substrate, translation initiation factor 2α (eIF2α). G-actin, whose essential role in eIF2α dephosphorylation is supported crystallographically, biochemically and genetically, aligns the catalytic and regulatory subunits, creating a composite surface that engages the N-terminal domain of eIF2α to position the distant phosphoserine-51 at the active site. Substrate residues that mediate affinity for the holophosphatase also make critical contacts with eIF2α kinases. Thus, a convergent process of higher-order substrate recognition specifies functionally antagonistic phosphorylation and dephosphorylation in the ISR. | ||||||
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 | 7nzm.cif.gz | 221.8 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7nzm.ent.gz | 174.6 KB | Display | PDB format |
PDBx/mmJSON format | 7nzm.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7nzm_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 7nzm_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 7nzm_validation.xml.gz | 51.4 KB | Display | |
Data in CIF | 7nzm_validation.cif.gz | 76.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/nz/7nzm ftp://data.pdbj.org/pub/pdb/validation_reports/nz/7nzm | HTTPS FTP |
-Related structure data
Related structure data | 12665MC 7nxvC 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
-Protein , 5 types, 5 molecules EBADC
#1: Protein | Mass: 21817.863 Da / Num. of mol.: 1 / Mutation: phosphorylated Ser51 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: EIF2S1, EIF2A / Production host: Escherichia coli (E. coli) / References: UniProt: P05198 |
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#2: Protein | Mass: 33647.621 Da / Num. of mol.: 1 / Mutation: D64A Source method: isolated from a genetically manipulated source Source: (gene. exp.) Oryctolagus cuniculus (rabbit) / Gene: PPP1CA, PPP1A / Production host: Escherichia coli (E. coli) References: UniProt: P62139, protein-serine/threonine phosphatase |
#3: Protein | Mass: 41862.613 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Oryctolagus cuniculus (rabbit) / References: UniProt: P68135 |
#4: Protein | Mass: 29092.574 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Bos taurus (cattle) / References: UniProt: P00639, deoxyribonuclease I |
#5: Protein | Mass: 49169.797 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human), (gene. exp.) Escherichia coli (strain K12) (bacteria) Gene: PPP1R15A, GADD34, malE, b4034, JW3994 / Strain: K12 / Production host: Escherichia coli (E. coli) / References: UniProt: O75807, UniProt: P0AEX9 |
-Sugars , 1 types, 1 molecules
#6: 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 |
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-Non-polymers , 2 types, 2 molecules
#7: Chemical | ChemComp-MN / |
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#8: Chemical | ChemComp-ATP / |
-Details
Has ligand of interest | Y |
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-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: pre-dephosphorylation complex of phosphorylated eIF2alpha_2-187 with trapped holophosphatase (PP1A_D64A/PPP1R15A_553-624/G-actin) Type: COMPLEX Details: One copy of each component was present in the complex: phosphorylated eIF2alpha_2-187, PP1A_D64A, PPP1R15A_553-624, G-actin and DNase I. The full complex was purified by size exclusion chromatography. Entity ID: #1-#5 / Source: MULTIPLE SOURCES | ||||||||||||||||||||||||||||||||
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Molecular weight | Value: 0.181 MDa / Experimental value: YES | ||||||||||||||||||||||||||||||||
Buffer solution | pH: 7.4 Details: 0.22mM Triton X-100 was added into the solution before plunging. | ||||||||||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | ||||||||||||||||||||||||||||||||
Specimen support | Details: current 25mA at Pelco EasiGLOW / Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R0.6/1 | ||||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-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: SPOT SCAN |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: -2800 nm / Nominal defocus min: -1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 46.84 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 4025 |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV / Phase plate: VOLTA PHASE PLATE |
-Processing
EM software |
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CTF correction | Details: Warp estimated the CTF parameters and passed them on to CryoSPARC to perform CTF correction. Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 132495 | ||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.96 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 60413 / Details: non-uniform refinement / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 47 / Protocol: OTHER / Space: REAL | ||||||||||||||||||||||||||||||||||||
Atomic model building |
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