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Yorodumi- EMDB-2666: Structure of SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-2666 | |||||||||
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Title | Structure of SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1 and UPF2 | |||||||||
Map data | Reconstruction of the SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1 and UPF2 | |||||||||
Sample |
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Keywords | NMD / SMG1 / SMG8 / SMG9 / UPF1 / UPF2 / PIKK / RNA degradation | |||||||||
Function / homology | Function and homology information double-stranded DNA helicase activity / supraspliceosomal complex / positive regulation of mRNA cis splicing, via spliceosome / RNA metabolic process / exon-exon junction complex / telomere maintenance via semi-conservative replication / positive regulation of mRNA catabolic process / cell cycle phase transition / diacylglycerol-dependent serine/threonine kinase activity / chromatoid body ...double-stranded DNA helicase activity / supraspliceosomal complex / positive regulation of mRNA cis splicing, via spliceosome / RNA metabolic process / exon-exon junction complex / telomere maintenance via semi-conservative replication / positive regulation of mRNA catabolic process / cell cycle phase transition / diacylglycerol-dependent serine/threonine kinase activity / chromatoid body / regulation of translational termination / histone mRNA catabolic process / eye development / 3'-UTR-mediated mRNA destabilization / regulation of protein kinase activity / regulation of telomere maintenance / nuclear-transcribed mRNA catabolic process, nonsense-mediated decay / DNA duplex unwinding / telomeric DNA binding / phosphatidylinositol phosphate biosynthetic process / nuclear-transcribed mRNA catabolic process / cellular response to interleukin-1 / Nonsense Mediated Decay (NMD) independent of the Exon Junction Complex (EJC) / animal organ regeneration / Nonsense Mediated Decay (NMD) enhanced by the Exon Junction Complex (EJC) / mRNA export from nucleus / liver development / helicase activity / P-body / brain development / Regulation of expression of SLITs and ROBOs / cytoplasmic ribonucleoprotein granule / heart development / peptidyl-serine phosphorylation / chromosome, telomeric region / DNA replication / cellular response to lipopolysaccharide / DNA helicase / protein autophosphorylation / in utero embryonic development / RNA helicase activity / non-specific serine/threonine protein kinase / protein kinase activity / RNA helicase / DNA repair / protein serine kinase activity / protein serine/threonine kinase activity / chromatin binding / DNA damage response / chromatin / negative regulation of apoptotic process / protein-containing complex binding / perinuclear region of cytoplasm / ATP hydrolysis activity / RNA binding / zinc ion binding / nucleoplasm / ATP binding / identical protein binding / nucleus / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / negative staining / Resolution: 19.4 Å | |||||||||
Authors | Melero R / Uchiyama A / Castano R / Kataoka N / Kurosawa H / Ohno S / Yamashita A / Llorca O | |||||||||
Citation | Journal: Structure / Year: 2014 Title: Structures of SMG1-UPFs complexes: SMG1 contributes to regulate UPF2-dependent activation of UPF1 in NMD. Authors: Roberto Melero / Akiko Uchiyama / Raquel Castaño / Naoyuki Kataoka / Hitomi Kurosawa / Shigeo Ohno / Akio Yamashita / Oscar Llorca / Abstract: SMG1, a PI3K-related kinase, plays a critical role in nonsense-mediated mRNA decay (NMD) in mammals. SMG1-mediated phosphorylation of the UPF1 helicase is an essential step during NMD initiation. ...SMG1, a PI3K-related kinase, plays a critical role in nonsense-mediated mRNA decay (NMD) in mammals. SMG1-mediated phosphorylation of the UPF1 helicase is an essential step during NMD initiation. Both SMG1 and UPF1 are presumably activated by UPF2, but this regulation is incompletely understood. Here we reveal that SMG1C (a complex containing SMG1, SMG8, and SMG9) contributes to regulate NMD by recruiting UPF1 and UPF2 to distinct sites in the vicinity of the kinase domain. UPF2 binds SMG1 in an UPF1-independent manner in vivo, and the SMG1C-UPF2 structure shows UPF2 recognizes the FRB domain, a region that regulates the related mTOR kinase. The molecular architectures of several SMG1C-UPFs complexes, obtained by combining electron microscopy with in vivo and in vitro interaction analyses, competition experiments, and mutations, suggest that UPF2 can be transferred to UPF1 within SMG1C, inducing UPF2-dependent conformational changes required to activate UPF1 within an SMG1C-UPF1-UPF2 complex. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_2666.map.gz | 9.3 MB | EMDB map data format | |
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Header (meta data) | emd-2666-v30.xml emd-2666.xml | 15.3 KB 15.3 KB | Display Display | EMDB header |
Images | emd_2666.png | 49.4 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-2666 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-2666 | HTTPS FTP |
-Validation report
Summary document | emd_2666_validation.pdf.gz | 202.9 KB | Display | EMDB validaton report |
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Full document | emd_2666_full_validation.pdf.gz | 202 KB | Display | |
Data in XML | emd_2666_validation.xml.gz | 5.8 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2666 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-2666 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_2666.map.gz / Format: CCP4 / Size: 11.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of the SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1 and UPF2 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 2.84 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1...
Entire | Name: SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1 and UPF2 |
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Components |
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-Supramolecule #1000: SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1...
Supramolecule | Name: SMG1C-UPF1-UPF2 complex, comprising SMG1 kinase, SMG8, SMG9, UPF1 and UPF2 type: sample / ID: 1000 / Number unique components: 5 |
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Molecular weight | Theoretical: 850 KDa |
-Macromolecule #1: Serine/threonine-protein kinase SMG1
Macromolecule | Name: Serine/threonine-protein kinase SMG1 / type: protein_or_peptide / ID: 1 / Name.synonym: SMG-1 / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Molecular weight | Theoretical: 410 KDa |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant cell: 293T cells |
Sequence | UniProtKB: Serine/threonine-protein kinase SMG1 GO: DNA repair, RNA metabolic process, nuclear-transcribed mRNA catabolic process, nonsense-mediated decay, ATP binding |
-Macromolecule #2: SMG8
Macromolecule | Name: SMG8 / type: protein_or_peptide / ID: 2 / Name.synonym: SMG-8 / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Molecular weight | Theoretical: 109 KDa |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant cell: 293T cells |
Sequence | UniProtKB: Nonsense-mediated mRNA decay factor SMG8 GO: nuclear-transcribed mRNA catabolic process, nonsense-mediated decay InterPro: Nonsense-mediated mRNA decay factor SMG8 |
-Macromolecule #3: SMG9
Macromolecule | Name: SMG9 / type: protein_or_peptide / ID: 3 / Name.synonym: SMG-9 / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Molecular weight | Theoretical: 60 KDa |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant cell: 293T cells |
Sequence | UniProtKB: Nonsense-mediated mRNA decay factor SMG9 GO: nuclear-transcribed mRNA catabolic process, nonsense-mediated decay InterPro: P-loop containing nucleoside triphosphate hydrolase, Nonsense-mediated mRNA decay factor SMG8/SMG9 |
-Macromolecule #4: UPF1
Macromolecule | Name: UPF1 / type: protein_or_peptide / ID: 4 / Name.synonym: RENT1_HUMAN / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Molecular weight | Theoretical: 120 KDa |
Recombinant expression | Organism: Homo sapiens (human) / Recombinant cell: 293T cells |
Sequence | UniProtKB: Regulator of nonsense transcripts 1 GO: nuclear-transcribed mRNA catabolic process, nonsense-mediated decay InterPro: P-loop containing nucleoside triphosphate hydrolase, RNA helicase UPF1, Cys/His rich zinc-binding domain |
-Macromolecule #5: UPF2
Macromolecule | Name: UPF2 / type: protein_or_peptide / ID: 5 / Name.synonym: RENT2_HUMAN / Recombinant expression: Yes |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human |
Molecular weight | Theoretical: 148 KDa |
Recombinant expression | Organism: Escherichia coli (E. coli) |
Sequence | UniProtKB: Regulator of nonsense transcripts 2 GO: nuclear-transcribed mRNA catabolic process, nonsense-mediated decay InterPro: Armadillo-type fold, INTERPRO: IPR016021, MIF4G-like, type 3, Up-frameshift suppressor 2, C-terminal |
-Experimental details
-Structure determination
Method | negative staining |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 0.01 mg/mL |
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Buffer | pH: 7.5 Details: 10 mM HEPES-KOH, 150 mM NaCl, 20% glycerol, 10 mM MgCl2 |
Staining | Type: NEGATIVE / Details: 1% uranyl formate |
Grid | Details: 400 mesh grid with thin carbon support, glow discharged |
Vitrification | Cryogen name: NONE / Instrument: OTHER |
-Electron microscopy
Microscope | JEOL 1230 |
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Alignment procedure | Legacy - Astigmatism: Objective lens astigmatism was corrected using a TVIPS F416 CMOS and the EM-MENU software (TVIPS) |
Date | Sep 12, 2012 |
Image recording | Category: CCD / Film or detector model: TVIPS TEMCAM-F416 (4k x 4k) / Digitization - Sampling interval: 15.6 µm / Number real images: 570 / Average electron dose: 15 e/Å2 Details: Using a TVIPS F416 CMOS and the EM-TOOLS software (TVIPS) Bits/pixel: 16 |
Electron beam | Acceleration voltage: 100 kV / Electron source: TUNGSTEN HAIRPIN |
Electron optics | Calibrated magnification: 54926 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.9 mm / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.5 µm / Nominal magnification: 40000 |
Sample stage | Specimen holder model: JEOL |
-Image processing
CTF correction | Details: Each micrograph using BSOFT |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 19.4 Å / Resolution method: OTHER / Software - Name: EMAN, EMAN2, Xmipp / Number images used: 32918 |
Final two d classification | Number classes: 490 |
-Atomic model buiding 1
Initial model | PDB ID: |
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Software | Name: Chimera |
Details | The structure was separately fitted using Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: Correlation coefficient |
-Atomic model buiding 2
Initial model | PDB ID: |
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Software | Name: Chimera |
Details | HEAT repeat regions from DNA-PKcs were separately fitted into SMG1 using Chimera |
Refinement | Space: REAL / Protocol: RIGID BODY FIT / Target criteria: Correlation coefficient |