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Yorodumi- PDB-8a9k: Cryo-EM structure of USP1-UAF1 bound to FANCI and mono-ubiquitina... -
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-Basic information
Entry | Database: PDB / ID: 8a9k | |||||||||
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Title | Cryo-EM structure of USP1-UAF1 bound to FANCI and mono-ubiquitinated FANCD2 with ML323 (consensus reconstruction) | |||||||||
Components |
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Keywords | HYDROLASE / Deubiquitinase / Complex / Enzyme-Substrate / Inhibitor | |||||||||
Function / homology | Function and homology information regulation of protein monoubiquitination / positive regulation of error-prone translesion synthesis / Signaling by cytosolic PDGFRA and PDGFRB fusion proteins / regulation of CD40 signaling pathway / regulation of regulatory T cell differentiation / monoubiquitinated protein deubiquitination / homologous chromosome pairing at meiosis / gamete generation / double-strand break repair involved in meiotic recombination / neuronal stem cell population maintenance ...regulation of protein monoubiquitination / positive regulation of error-prone translesion synthesis / Signaling by cytosolic PDGFRA and PDGFRB fusion proteins / regulation of CD40 signaling pathway / regulation of regulatory T cell differentiation / monoubiquitinated protein deubiquitination / homologous chromosome pairing at meiosis / gamete generation / double-strand break repair involved in meiotic recombination / neuronal stem cell population maintenance / brain morphogenesis / deubiquitinase activator activity / DNA repair complex / mitotic intra-S DNA damage checkpoint signaling / skeletal system morphogenesis / skin development / protein deubiquitination / seminiferous tubule development / homeostasis of number of cells / positive regulation of double-strand break repair via homologous recombination / single fertilization / embryonic organ development / interstrand cross-link repair / regulation of DNA repair / DNA polymerase binding / response to UV / condensed chromosome / Maturation of protein E / Maturation of protein E / ER Quality Control Compartment (ERQC) / Myoclonic epilepsy of Lafora / IRAK2 mediated activation of TAK1 complex / Alpha-protein kinase 1 signaling pathway / FLT3 signaling by CBL mutants / IRAK1 recruits IKK complex / IRAK1 recruits IKK complex upon TLR7/8 or 9 stimulation / Prevention of phagosomal-lysosomal fusion / Glycogen synthesis / IRAK2 mediated activation of TAK1 complex upon TLR7/8 or 9 stimulation / Regulation of TBK1, IKKε (IKBKE)-mediated activation of IRF3, IRF7 / Endosomal Sorting Complex Required For Transport (ESCRT) / TICAM1,TRAF6-dependent induction of TAK1 complex / Membrane binding and targetting of GAG proteins / Regulation of TBK1, IKKε-mediated activation of IRF3, IRF7 upon TLR3 ligation / Negative regulation of FLT3 / Constitutive Signaling by NOTCH1 HD Domain Mutants / PTK6 Regulates RTKs and Their Effectors AKT1 and DOK1 / Regulation of FZD by ubiquitination / TICAM1-dependent activation of IRF3/IRF7 / NOTCH2 Activation and Transmission of Signal to the Nucleus / p75NTR recruits signalling complexes / APC/C:Cdc20 mediated degradation of Cyclin B / Downregulation of ERBB4 signaling / TRAF6-mediated induction of TAK1 complex within TLR4 complex / TRAF6 mediated IRF7 activation in TLR7/8 or 9 signaling / APC-Cdc20 mediated degradation of Nek2A / Regulation of innate immune responses to cytosolic DNA / NF-kB is activated and signals survival / InlA-mediated entry of Listeria monocytogenes into host cells / Regulation of pyruvate metabolism / VLDLR internalisation and degradation / Downregulation of ERBB2:ERBB3 signaling / NRIF signals cell death from the nucleus / Pexophagy / Activated NOTCH1 Transmits Signal to the Nucleus / Regulation of PTEN localization / Regulation of BACH1 activity / Synthesis of active ubiquitin: roles of E1 and E2 enzymes / TICAM1, RIP1-mediated IKK complex recruitment / Translesion synthesis by REV1 / MAP3K8 (TPL2)-dependent MAPK1/3 activation / Translesion synthesis by POLK / Downregulation of TGF-beta receptor signaling / Activation of IRF3, IRF7 mediated by TBK1, IKKε (IKBKE) / Translesion synthesis by POLI / PINK1-PRKN Mediated Mitophagy / IKK complex recruitment mediated by RIP1 / Regulation of activated PAK-2p34 by proteasome mediated degradation / JNK (c-Jun kinases) phosphorylation and activation mediated by activated human TAK1 / Gap-filling DNA repair synthesis and ligation in GG-NER / Josephin domain DUBs / InlB-mediated entry of Listeria monocytogenes into host cell / positive regulation of epithelial cell proliferation / TGF-beta receptor signaling in EMT (epithelial to mesenchymal transition) / ubiquitin binding / positive regulation of protein ubiquitination / TNFR1-induced NF-kappa-B signaling pathway / N-glycan trimming in the ER and Calnexin/Calreticulin cycle / Autodegradation of Cdh1 by Cdh1:APC/C / skeletal system development / APC/C:Cdc20 mediated degradation of Securin / Regulation of NF-kappa B signaling / SCF-beta-TrCP mediated degradation of Emi1 / Asymmetric localization of PCP proteins / TCF dependent signaling in response to WNT / NIK-->noncanonical NF-kB signaling / Ubiquitin-dependent degradation of Cyclin D / AUF1 (hnRNP D0) binds and destabilizes mRNA / activated TAK1 mediates p38 MAPK activation / TNFR2 non-canonical NF-kB pathway Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) synthetic construct (others) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.85 Å | |||||||||
Authors | Rennie, M.L. / Walden, H. | |||||||||
Funding support | European Union, United Kingdom, 2items
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Citation | Journal: Sci Adv / Year: 2022 Title: Cryo-EM reveals a mechanism of USP1 inhibition through a cryptic binding site. Authors: Martin L Rennie / Connor Arkinson / Viduth K Chaugule / Helen Walden / Abstract: Repair of DNA damage is critical to genomic integrity and frequently disrupted in cancers. Ubiquitin-specific protease 1 (USP1), a nucleus-localized deubiquitinase, lies at the interface of multiple ...Repair of DNA damage is critical to genomic integrity and frequently disrupted in cancers. Ubiquitin-specific protease 1 (USP1), a nucleus-localized deubiquitinase, lies at the interface of multiple DNA repair pathways and is a promising drug target for certain cancers. Although multiple inhibitors of this enzyme, including one in phase 1 clinical trials, have been established, their binding mode is unknown. Here, we use cryo-electron microscopy to study an assembled enzyme-substrate-inhibitor complex of USP1 and the well-established inhibitor, ML323. Achieving 2.5-Å resolution, with and without ML323, we find an unusual binding mode in which the inhibitor disrupts part of the hydrophobic core of USP1. The consequent conformational changes in the secondary structure lead to subtle rearrangements in the active site that underlie the mechanism of inhibition. These structures provide a platform for structure-based drug design targeting USP1. | |||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8a9k.cif.gz | 563.5 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8a9k.ent.gz | 437.7 KB | Display | PDB format |
PDBx/mmJSON format | 8a9k.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 8a9k_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 8a9k_full_validation.pdf.gz | 1.4 MB | Display | |
Data in XML | 8a9k_validation.xml.gz | 87.3 KB | Display | |
Data in CIF | 8a9k_validation.cif.gz | 130.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/a9/8a9k ftp://data.pdbj.org/pub/pdb/validation_reports/a9/8a9k | HTTPS FTP |
-Related structure data
Related structure data | 14722MC 7zh3C 7zh4C 8a9jC C: citing same article (ref.) M: map data used to model this data |
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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
-Fanconi anemia group ... , 2 types, 2 molecules AB
#1: Protein | Mass: 150459.125 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: FANCI, KIAA1794 / Cell line (production host): Sf21 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9NVI1 |
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#2: Protein | Mass: 164623.828 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: FANCD2, FACD / Cell line (production host): Sf21 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q9BXW9 |
-Protein , 3 types, 3 molecules CDE
#3: Protein | Mass: 8875.125 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: UBC / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P0CG48 |
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#4: Protein | Mass: 88390.273 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: USP1 / Cell line (production host): Sf21 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: O94782, ubiquitinyl hydrolase 1 |
#5: Protein | Mass: 78300.656 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: WDR48, KIAA1449, UAF1 / Cell line (production host): Sf21 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q8TAF3 |
-DNA chain , 1 types, 2 molecules ST
#6: DNA chain | Mass: 5177.820 Da / Num. of mol.: 2 / Source method: obtained synthetically Details: Actual sequence: TGATCAGAGGTCATTTGAATTCATGGCTTCGAGCTTCATGTAGAGTCGACGGTGCTGGGAT Source: (synth.) synthetic construct (others) |
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-Non-polymers , 2 types, 2 molecules
#7: Chemical | ChemComp-ZN / |
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#8: Chemical | ChemComp-JDA / |
-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: USP1(C90S)-UAF1 bound to FANCI and mono-ubiquitinated FANCD2 with dsDNA and ML323 Type: COMPLEX / Entity ID: #1-#6 / Source: RECOMBINANT |
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Molecular weight | Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Spodoptera frugiperda (fall armyworm) |
Buffer solution | pH: 8 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES Details: 9.3 uM USP1-UAF1, 1.8 uM FANCI-FANCD2Ub, 2.2 uM dsDNA (61 base-pairs), 18 uM ML323 |
Specimen support | Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 |
Vitrification | Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 288 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: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal defocus max: 3000 nm / Nominal defocus min: 1000 nm |
Image recording | Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
-Processing
Software | Name: PHENIX / Version: 1.19.2_4158: / Classification: refinement | ||||||||||||||||||||||||||||
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EM software |
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Image processing | Details: 2x binning | ||||||||||||||||||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 6716868 | ||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.85 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 570816 / Algorithm: BACK PROJECTION / Symmetry type: POINT | ||||||||||||||||||||||||||||
Atomic model building | B value: 95.3 / Space: REAL | ||||||||||||||||||||||||||||
Refine LS restraints |
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