+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-13443 | |||||||||
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Title | DNA-PK in complex with ATPgammaS | |||||||||
Map data | ||||||||||
Sample |
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Function / homology | Function and homology information positive regulation of platelet formation / T cell receptor V(D)J recombination / pro-B cell differentiation / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase activity / histone H2AXS139 kinase activity / DNA-dependent protein kinase complex / immature B cell differentiation / DNA-dependent protein kinase-DNA ligase 4 complex ...positive regulation of platelet formation / T cell receptor V(D)J recombination / pro-B cell differentiation / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase activity / histone H2AXS139 kinase activity / DNA-dependent protein kinase complex / immature B cell differentiation / DNA-dependent protein kinase-DNA ligase 4 complex / immunoglobulin V(D)J recombination / nonhomologous end joining complex / regulation of smooth muscle cell proliferation / double-strand break repair via alternative nonhomologous end joining / Cytosolic sensors of pathogen-associated DNA / regulation of epithelial cell proliferation / IRF3-mediated induction of type I IFN / telomere capping / U3 snoRNA binding / regulation of hematopoietic stem cell differentiation / maturation of 5.8S rRNA / T cell lineage commitment / negative regulation of cGAS/STING signaling pathway / B cell lineage commitment / positive regulation of double-strand break repair via nonhomologous end joining / ectopic germ cell programmed cell death / somitogenesis / mitotic G1 DNA damage checkpoint signaling / telomere maintenance / activation of innate immune response / small-subunit processome / positive regulation of translation / negative regulation of protein phosphorylation / positive regulation of erythrocyte differentiation / protein-DNA complex / response to gamma radiation / Nonhomologous End-Joining (NHEJ) / brain development / peptidyl-threonine phosphorylation / protein modification process / protein destabilization / regulation of circadian rhythm / double-strand break repair via nonhomologous end joining / cellular response to insulin stimulus / rhythmic process / intrinsic apoptotic signaling pathway in response to DNA damage / double-strand break repair / E3 ubiquitin ligases ubiquitinate target proteins / T cell differentiation in thymus / heart development / double-stranded DNA binding / peptidyl-serine phosphorylation / transcription regulator complex / RNA polymerase II-specific DNA-binding transcription factor binding / chromosome, telomeric region / non-specific serine/threonine protein kinase / protein kinase activity / positive regulation of apoptotic process / protein domain specific binding / protein phosphorylation / protein serine kinase activity / innate immune response / protein serine/threonine kinase activity / DNA damage response / chromatin / nucleolus / negative regulation of apoptotic process / enzyme binding / positive regulation of transcription by RNA polymerase II / protein-containing complex / RNA binding / nucleoplasm / ATP binding / membrane / nucleus / cytosol Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.3 Å | |||||||||
Authors | Liang S / Blundell TL | |||||||||
Funding support | United Kingdom, 1 items
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Citation | Journal: Nature / Year: 2022 Title: Structural insights into inhibitor regulation of the DNA repair protein DNA-PKcs. Authors: Shikang Liang / Sherine E Thomas / Amanda K Chaplin / Steven W Hardwick / Dimitri Y Chirgadze / Tom L Blundell / Abstract: The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) has a central role in non-homologous end joining, one of the two main pathways that detect and repair DNA double-strand breaks (DSBs) in ...The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) has a central role in non-homologous end joining, one of the two main pathways that detect and repair DNA double-strand breaks (DSBs) in humans. DNA-PKcs is of great importance in repairing pathological DSBs, making DNA-PKcs inhibitors attractive therapeutic agents for cancer in combination with DSB-inducing radiotherapy and chemotherapy. Many of the selective inhibitors of DNA-PKcs that have been developed exhibit potential as treatment for various cancers. Here we report cryo-electron microscopy (cryo-EM) structures of human DNA-PKcs natively purified from HeLa cell nuclear extracts, in complex with adenosine-5'-(γ-thio)-triphosphate (ATPγS) and four inhibitors (wortmannin, NU7441, AZD7648 and M3814), including drug candidates undergoing clinical trials. The structures reveal molecular details of ATP binding at the active site before catalysis and provide insights into the modes of action and specificities of the competitive inhibitors. Of note, binding of the ligands causes movement of the PIKK regulatory domain (PRD), revealing a connection between the p-loop and PRD conformations. Electrophoretic mobility shift assay and cryo-EM studies on the DNA-dependent protein kinase holoenzyme further show that ligand binding does not have a negative allosteric or inhibitory effect on assembly of the holoenzyme complex and that inhibitors function through direct competition with ATP. Overall, the structures described in this study should greatly assist future efforts in rational drug design targeting DNA-PKcs, demonstrating the potential of cryo-EM in structure-guided drug development for large and challenging targets. | |||||||||
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_13443.map.gz | 154.7 MB | EMDB map data format | |
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Header (meta data) | emd-13443-v30.xml emd-13443.xml | 7.5 KB 7.5 KB | Display Display | EMDB header |
Images | emd_13443.png | 35.3 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-13443 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-13443 | 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_13443.map.gz / Format: CCP4 / Size: 163.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.304 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 : Holoenzyme of DNA-PK, including DNA-PKcs and Ku70/80 heterodimer,...
Entire | Name: Holoenzyme of DNA-PK, including DNA-PKcs and Ku70/80 heterodimer, in complex with ATPgammaS |
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Components |
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-Supramolecule #1: Holoenzyme of DNA-PK, including DNA-PKcs and Ku70/80 heterodimer,...
Supramolecule | Name: Holoenzyme of DNA-PK, including DNA-PKcs and Ku70/80 heterodimer, in complex with ATPgammaS type: complex / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Homo sapiens (human) |
Molecular weight | Experimental: 660 KDa |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 47.9 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Initial angle assignment | Type: MAXIMUM LIKELIHOOD |
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Final angle assignment | Type: MAXIMUM LIKELIHOOD |
Final reconstruction | Resolution.type: BY AUTHOR / Resolution: 4.3 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 18758 |