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Yorodumi- PDB-8pn9: Structure of human oligosaccharyltransferase OST-A complex bound ... -
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-Basic information
Entry | Database: PDB / ID: 8pn9 | ||||||
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Title | Structure of human oligosaccharyltransferase OST-A complex bound to NGI-1 | ||||||
Components |
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Keywords | TRANSFERASE / N-glycosylation / OST-A complex / NGI-1 inhibitor | ||||||
Function / homology | Function and homology information oligosaccharyltransferase complex binding / oligosaccharyltransferase I complex / oligosaccharyltransferase III complex / membrane-bounded organelle / Asparagine N-linked glycosylation / co-translational protein modification / oligosaccharyltransferase complex / dolichyl-diphosphooligosaccharide-protein glycotransferase / dolichyl-diphosphooligosaccharide-protein glycotransferase activity / protein N-linked glycosylation via asparagine ...oligosaccharyltransferase complex binding / oligosaccharyltransferase I complex / oligosaccharyltransferase III complex / membrane-bounded organelle / Asparagine N-linked glycosylation / co-translational protein modification / oligosaccharyltransferase complex / dolichyl-diphosphooligosaccharide-protein glycotransferase / dolichyl-diphosphooligosaccharide-protein glycotransferase activity / protein N-linked glycosylation via asparagine / protein N-linked glycosylation / epithelial cell apoptotic process / azurophil granule membrane / protein glycosylation / blastocyst development / Advanced glycosylation endproduct receptor signaling / SRP-dependent cotranslational protein targeting to membrane / rough endoplasmic reticulum / enzyme activator activity / T cell activation / response to endoplasmic reticulum stress / post-translational protein modification / response to cytokine / protein modification process / regulation of protein stability / melanosome / Maturation of spike protein / nuclear body / inflammatory response / intracellular membrane-bounded organelle / apoptotic process / Neutrophil degranulation / endoplasmic reticulum membrane / negative regulation of apoptotic process / endoplasmic reticulum / protein-containing complex / RNA binding / membrane / metal ion binding / plasma membrane / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.61 Å | ||||||
Authors | Ramirez, A.S. / Kowal, J. / Locher, K.P. | ||||||
Funding support | Switzerland, 1items
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Citation | Journal: Cell / Year: 2024 Title: Positive selection CRISPR screens reveal a druggable pocket in an oligosaccharyltransferase required for inflammatory signaling to NF-κB. Authors: Benjamin L Lampson / Ana S Ramίrez / Marta Baro / Lixia He / Mudra Hegde / Vidyasagar Koduri / Jamie L Pfaff / Ruth E Hanna / Julia Kowal / Nitin H Shirole / Yanfeng He / John G Doench / ...Authors: Benjamin L Lampson / Ana S Ramίrez / Marta Baro / Lixia He / Mudra Hegde / Vidyasagar Koduri / Jamie L Pfaff / Ruth E Hanna / Julia Kowal / Nitin H Shirole / Yanfeng He / John G Doench / Joseph N Contessa / Kaspar P Locher / William G Kaelin / Abstract: Nuclear factor κB (NF-κB) plays roles in various diseases. Many inflammatory signals, such as circulating lipopolysaccharides (LPSs), activate NF-κB via specific receptors. Using whole-genome ...Nuclear factor κB (NF-κB) plays roles in various diseases. Many inflammatory signals, such as circulating lipopolysaccharides (LPSs), activate NF-κB via specific receptors. Using whole-genome CRISPR-Cas9 screens of LPS-treated cells that express an NF-κB-driven suicide gene, we discovered that the LPS receptor Toll-like receptor 4 (TLR4) is specifically dependent on the oligosaccharyltransferase complex OST-A for N-glycosylation and cell-surface localization. The tool compound NGI-1 inhibits OST complexes in vivo, but the underlying molecular mechanism remained unknown. We did a CRISPR base-editor screen for NGI-1-resistant variants of STT3A, the catalytic subunit of OST-A. These variants, in conjunction with cryoelectron microscopy studies, revealed that NGI-1 binds the catalytic site of STT3A, where it traps a molecule of the donor substrate dolichyl-PP-GlcNAc-Man-Glc, suggesting an uncompetitive inhibition mechanism. Our results provide a rationale for and an initial step toward the development of STT3A-specific inhibitors and illustrate the power of contemporaneous base-editor and structural studies to define drug mechanism of action. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8pn9.cif.gz | 423.2 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8pn9.ent.gz | 331.6 KB | Display | PDB format |
PDBx/mmJSON format | 8pn9.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8pn9_validation.pdf.gz | 2.4 MB | Display | wwPDB validaton report |
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Full document | 8pn9_full_validation.pdf.gz | 2.4 MB | Display | |
Data in XML | 8pn9_validation.xml.gz | 81.2 KB | Display | |
Data in CIF | 8pn9_validation.cif.gz | 114.6 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/pn/8pn9 ftp://data.pdbj.org/pub/pdb/validation_reports/pn/8pn9 | HTTPS FTP |
-Related structure data
Related structure data | 17779MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Dolichyl-diphosphooligosaccharide--protein glycosyltransferase subunit ... , 5 types, 5 molecules ABDEF
#1: Protein | Mass: 80607.320 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: STT3A, ITM1, TMC / Production host: Homo sapiens (human) References: UniProt: P46977, dolichyl-diphosphooligosaccharide-protein glycotransferase |
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#2: Protein/peptide | Mass: 4196.004 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: OST4 / Production host: Homo sapiens (human) / References: UniProt: P0C6T2 |
#4: Protein | Mass: 12503.631 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: DAD1 / Production host: Homo sapiens (human) / References: UniProt: P61803 |
#5: Protein | Mass: 68656.156 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RPN1 / Production host: Homo sapiens (human) / References: UniProt: P04843 |
#6: Protein | Mass: 69347.508 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: RPN2 / Production host: Homo sapiens (human) / References: UniProt: P04844 |
-Protein , 3 types, 3 molecules CHG
#3: Protein | Mass: 9083.804 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: TMEM258, C11orf10, HSPC005 / Production host: Homo sapiens (human) / References: UniProt: P61165 |
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#7: Protein | Mass: 16844.215 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: OSTC, DC2, HDCMD45P, HSPC307 / Production host: Homo sapiens (human) / References: UniProt: Q9NRP0 |
#8: Protein | Mass: 50293.977 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: DDOST, KIAA0115, OST48, OK/SW-cl.45 / Production host: Homo sapiens (human) / References: UniProt: P39656 |
-Sugars , 4 types, 4 molecules
#9: 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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#10: Polysaccharide | alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D- ...alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)-alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source |
#11: Polysaccharide | alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]alpha-D-mannopyranose-(1-6)-[alpha-D- ...alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)]alpha-D-mannopyranose-(1-6)-[alpha-D-mannopyranose-(1-3)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose Source method: isolated from a genetically manipulated source |
#12: Polysaccharide | alpha-D-glucopyranose-(1-2)-alpha-D-glucopyranose-(1-3)-alpha-D-glucopyranose-(1-3)-alpha-D- ...alpha-D-glucopyranose-(1-2)-alpha-D-glucopyranose-(1-3)-alpha-D-glucopyranose-(1-3)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[alpha-D-mannopyranose-(1-6)-alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-alpha-D-glucopyranose Type: oligosaccharide / Mass: 1883.668 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source |
-Non-polymers , 6 types, 10 molecules
#13: Chemical | ChemComp-KZB / ( #14: Chemical | #15: Chemical | ChemComp-MN / | #16: Chemical | ChemComp-OTP / ( | #17: Chemical | ChemComp-ZXT / | Mass: 394.512 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C17H22N4O3S2 / Feature type: SUBJECT OF INVESTIGATION #18: Water | ChemComp-HOH / | |
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-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: OLIGOSACCHARYLTRANSFERASE COMPLEX OST-A WITH BOUND NGI-1 INHIBITOR Type: COMPLEX / Entity ID: #2-#5, #7 / Source: NATURAL |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE-PROPANE |
-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: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 2400 nm / Nominal defocus min: 600 nm |
Image recording | Electron dose: 54.2 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION |
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3D reconstruction | Resolution: 3.61 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 101699 / Symmetry type: POINT |