+Open data
-Basic information
Entry | Database: PDB / ID: 7zwa | ||||||
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Title | CryoEM structure of Ku heterodimer bound to DNA and PAXX | ||||||
Components |
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Keywords | DNA BINDING PROTEIN / NHEJ / Ku70 / Ku80 / DNA damage / PAXX | ||||||
Function / homology | Function and homology information Ku70:Ku80 complex / negative regulation of t-circle formation / DNA end binding / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase complex / DNA-dependent protein kinase-DNA ligase 4 complex / cellular response to X-ray / nonhomologous end joining complex / DNA ligation ...Ku70:Ku80 complex / negative regulation of t-circle formation / DNA end binding / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase complex / DNA-dependent protein kinase-DNA ligase 4 complex / cellular response to X-ray / nonhomologous end joining complex / DNA ligation / regulation of smooth muscle cell proliferation / nuclear telomere cap complex / Cytosolic sensors of pathogen-associated DNA / double-strand break repair via classical nonhomologous end joining / IRF3-mediated induction of type I IFN / recombinational repair / regulation of telomere maintenance / U3 snoRNA binding / positive regulation of neurogenesis / cellular response to fatty acid / hematopoietic stem cell proliferation / protein localization to chromosome, telomeric region / cellular hyperosmotic salinity response / telomeric DNA binding / positive regulation of catalytic activity / 2-LTR circle formation / site of DNA damage / Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases / 5'-deoxyribose-5-phosphate lyase activity / hematopoietic stem cell differentiation / positive regulation of protein kinase activity / ATP-dependent activity, acting on DNA / DNA polymerase binding / DNA helicase activity / : / enzyme activator activity / positive regulation of telomere maintenance via telomerase / activation of innate immune response / telomere maintenance / cyclin binding / neurogenesis / cellular response to leukemia inhibitory factor / protein-DNA complex / small-subunit processome / Nonhomologous End-Joining (NHEJ) / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / cellular response to gamma radiation / double-strand break repair via nonhomologous end joining / double-strand break repair / site of double-strand break / scaffold protein binding / double-stranded DNA binding / secretory granule lumen / DNA recombination / ficolin-1-rich granule lumen / transcription regulator complex / chromosome, telomeric region / damaged DNA binding / molecular adaptor activity / transcription cis-regulatory region binding / ribonucleoprotein complex / response to xenobiotic stimulus / innate immune response / negative regulation of DNA-templated transcription / ubiquitin protein ligase binding / DNA damage response / Neutrophil degranulation / protein-containing complex binding / nucleolus / positive regulation of DNA-templated transcription / protein homodimerization activity / ATP hydrolysis activity / positive regulation of transcription by RNA polymerase II / protein-containing complex / DNA binding / RNA binding / extracellular region / nucleoplasm / ATP binding / identical protein binding / membrane / nucleus / plasma membrane / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | Homo sapiens (human) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.8 Å | ||||||
Authors | Hardwick, S.W. / Kefala-Stavridi, A. / Chirgadze, D.Y. / Blundell, T.L. / Chaplin, A.K. | ||||||
Funding support | United Kingdom, 1items
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Citation | Journal: Sci Adv / Year: 2023 Title: PAXX binding to the NHEJ machinery explains functional redundancy with XLF. Authors: Murielle Seif-El-Dahan / Antonia Kefala-Stavridi / Philippe Frit / Steven W Hardwick / Dima Y Chirgadze / Taiana Maia De Oliviera / Jessica Andreani / Sébastien Britton / Nadia Barboule / ...Authors: Murielle Seif-El-Dahan / Antonia Kefala-Stavridi / Philippe Frit / Steven W Hardwick / Dima Y Chirgadze / Taiana Maia De Oliviera / Jessica Andreani / Sébastien Britton / Nadia Barboule / Madeleine Bossaert / Arun Prasad Pandurangan / Katheryn Meek / Tom L Blundell / Virginie Ropars / Patrick Calsou / Jean-Baptiste Charbonnier / Amanda K Chaplin / Abstract: Nonhomologous end joining is a critical mechanism that repairs DNA double-strand breaks in human cells. In this work, we address the structural and functional role of the accessory protein PAXX ...Nonhomologous end joining is a critical mechanism that repairs DNA double-strand breaks in human cells. In this work, we address the structural and functional role of the accessory protein PAXX [paralog of x-ray repair cross-complementing protein 4 (XRCC4) and XRCC4-like factor (XLF)] in this mechanism. Here, we report high-resolution cryo-electron microscopy (cryo-EM) and x-ray crystallography structures of the PAXX C-terminal Ku-binding motif bound to Ku70/80 and cryo-EM structures of PAXX bound to two alternate DNA-dependent protein kinase (DNA-PK) end-bridging dimers, mediated by either Ku80 or XLF. We identify residues critical for the Ku70/PAXX interaction in vitro and in cells. We demonstrate that PAXX and XLF can bind simultaneously to the Ku heterodimer and act as structural bridges in alternate forms of DNA-PK dimers. Last, we show that engagement of both proteins provides a complementary advantage for DNA end synapsis and end joining in cells. | ||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 7zwa.cif.gz | 301.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7zwa.ent.gz | 187.4 KB | Display | PDB format |
PDBx/mmJSON format | 7zwa.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7zwa_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 7zwa_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | 7zwa_validation.xml.gz | 47.8 KB | Display | |
Data in CIF | 7zwa_validation.cif.gz | 70.2 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/zw/7zwa ftp://data.pdbj.org/pub/pdb/validation_reports/zw/7zwa | HTTPS FTP |
-Related structure data
Related structure data | 14995MC 7zygC 8ascC 8bh3C 8bhvC 8bhyC 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
-X-ray repair cross-complementing protein ... , 2 types, 2 molecules AB
#1: Protein | Mass: 69945.039 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: XRCC6, G22P1 / Production host: Spodoptera frugiperda (fall armyworm) References: UniProt: P12956, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement, Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases |
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#2: Protein | Mass: 82812.438 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: XRCC5, G22P2 / Production host: Spodoptera frugiperda (fall armyworm) References: UniProt: P13010, Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement |
-DNA chain , 2 types, 2 molecules DE
#4: DNA chain | Mass: 4543.972 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) |
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#5: DNA chain | Mass: 4962.240 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Homo sapiens (human) |
-Protein / Non-polymers , 2 types, 2 molecules C
#3: Protein | Mass: 21663.498 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: PAXX, C9orf142, XLS / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9BUH6 |
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#6: Chemical | ChemComp-PO4 / |
-Details
Has ligand of interest | N |
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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: Ku70/80 heterodimer bound to DNA and PAXX / Type: ORGANELLE OR CELLULAR COMPONENT / Entity ID: #1-#5 / Source: MULTIPLE SOURCES |
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Molecular weight | Value: 0.15 MDa / Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Spodoptera frugiperda (fall armyworm) |
Buffer solution | pH: 8 |
Specimen | Conc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
-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 magnification: 130000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 1.25 sec. / Electron dose: 44.39 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 6924 |
-Processing
Software |
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2143882 | |||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 161215 / Symmetry type: POINT | |||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | |||||||||||||||||||||||||||
Displacement parameters | Biso mean: 135.26 Å2 | |||||||||||||||||||||||||||
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