+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28733 | |||||||||
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Title | NHEJ Long-range complex with ATP | |||||||||
Map data | ATPAdenosine triphosphate | |||||||||
Sample |
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Keywords | Complex / Kinase / NHEJ / DNA BINDING PROTEIN / DNA BINDING PROTEIN-DNA complex | |||||||||
Function / homology | Function and homology information DNA ligation involved in DNA recombination / FHA domain binding / positive regulation of chromosome organization / positive regulation of ligase activity / DNA ligase IV complex / DNA ligation involved in DNA repair / DNA ligase activity / Ku70:Ku80 complex / DN2 thymocyte differentiation / negative regulation of t-circle formation ...DNA ligation involved in DNA recombination / FHA domain binding / positive regulation of chromosome organization / positive regulation of ligase activity / DNA ligase IV complex / DNA ligation involved in DNA repair / DNA ligase activity / Ku70:Ku80 complex / DN2 thymocyte differentiation / negative regulation of t-circle formation / positive regulation of platelet formation / DNA ligase (ATP) / DNA end binding / T cell receptor V(D)J recombination / pro-B cell differentiation / DNA ligase (ATP) activity / 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 / nucleotide-excision repair, DNA gap filling / DNA-dependent protein kinase-DNA ligase 4 complex / single strand break repair / cellular response to X-ray / immunoglobulin V(D)J recombination / nonhomologous end joining complex / protein localization to site of double-strand break / DNA ligation / nuclear telomere cap complex / regulation of smooth muscle cell proliferation / V(D)J recombination / double-strand break repair via alternative nonhomologous end joining / isotype switching / Cytosolic sensors of pathogen-associated DNA / double-strand break repair via classical nonhomologous end joining / regulation of epithelial cell proliferation / IRF3-mediated induction of type I IFN / telomere capping / recombinational repair / regulation of telomere maintenance / U3 snoRNA binding / regulation of hematopoietic stem cell differentiation / positive regulation of neurogenesis / protein localization to chromosome, telomeric region / cellular response to fatty acid / hematopoietic stem cell proliferation / cellular hyperosmotic salinity response / response to ionizing radiation / maturation of 5.8S rRNA / T cell lineage commitment / negative regulation of cGAS/STING signaling pathway / telomeric DNA binding / DNA biosynthetic process / positive regulation of catalytic activity / B cell lineage commitment / cellular response to lithium ion / 2-LTR circle formation / positive regulation of double-strand break repair via nonhomologous end joining / somatic stem cell population maintenance / site of DNA damage / ligase activity / Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases / enzyme activator activity / T cell differentiation / response to X-ray / 5'-deoxyribose-5-phosphate lyase activity / chromosome organization / hematopoietic stem cell differentiation / positive regulation of protein kinase activity / ectopic germ cell programmed cell death / ATP-dependent activity, acting on DNA / somitogenesis / DNA polymerase binding / SUMOylation of DNA damage response and repair proteins / condensed chromosome / positive regulation of telomerase activity / mitotic G1 DNA damage checkpoint signaling / positive regulation of telomere maintenance via telomerase / DNA helicase activity / activation of innate immune response / telomere maintenance / cellular response to leukemia inhibitory factor / neurogenesis / cyclin binding / small-subunit processome / B cell differentiation / positive regulation of translation / positive regulation of erythrocyte differentiation / negative regulation of protein phosphorylation / central nervous system development / stem cell proliferation / cellular response to ionizing radiation / protein-DNA complex / response to gamma radiation / Nonhomologous End-Joining (NHEJ) / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / brain development / peptidyl-threonine phosphorylation Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) / human (human) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 8.9 Å | |||||||||
Authors | Chen S / He Y | |||||||||
Funding support | United States, 2 items
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Citation | Journal: Sci Adv / Year: 2023 Title: Cryo-EM visualization of DNA-PKcs structural intermediates in NHEJ. Authors: Siyu Chen / Alex Vogt / Linda Lee / Tasmin Naila / Ryan McKeown / Alan E Tomkinson / Susan P Lees-Miller / Yuan He / Abstract: DNA double-strand breaks (DSBs), one of the most cytotoxic forms of DNA damage, can be repaired by the tightly regulated nonhomologous end joining (NHEJ) machinery (Stinson and Loparo and Zhao ). ...DNA double-strand breaks (DSBs), one of the most cytotoxic forms of DNA damage, can be repaired by the tightly regulated nonhomologous end joining (NHEJ) machinery (Stinson and Loparo and Zhao ). Core NHEJ factors form an initial long-range (LR) synaptic complex that transitions into a DNA-PKcs (DNA-dependent protein kinase, catalytic subunit)-free, short-range state to align the DSB ends (Chen ). Using single-particle cryo-electron microscopy, we have visualized three additional key NHEJ complexes representing different transition states, with DNA-PKcs adopting distinct dimeric conformations within each of them. Upon DNA-PKcs autophosphorylation, the LR complex undergoes a substantial conformational change, with both Ku and DNA-PKcs rotating outward to promote DNA break exposure and DNA-PKcs dissociation. We also captured a dimeric state of catalytically inactive DNA-PKcs, which resembles structures of other PIKK (Phosphatidylinositol 3-kinase-related kinase) family kinases, revealing a model of the full regulatory cycle of DNA-PKcs during NHEJ. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_28733.map.gz | 49.1 MB | EMDB map data format | |
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Header (meta data) | emd-28733-v30.xml emd-28733.xml | 33.9 KB 33.9 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_28733_fsc.xml | 8.6 KB | Display | FSC data file |
Images | emd_28733.png | 56.8 KB | ||
Others | emd_28733_half_map_1.map.gz emd_28733_half_map_2.map.gz | 39.5 MB 39.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28733 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28733 | HTTPS FTP |
-Related structure data
Related structure data | 8ezbMC 8ez9C 8ezaC C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_28733.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | ATP | ||||||||||||||||||||
Voxel size | X=Y=Z: 2.112 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: half1
File | emd_28733_half_map_1.map | ||||||||||||
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Annotation | half1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: half2
File | emd_28733_half_map_2.map | ||||||||||||
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Annotation | half2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : NHEJ Long-range complex with ATP
+Supramolecule #1: NHEJ Long-range complex with ATP
+Macromolecule #1: X-ray repair cross-complementing protein 6
+Macromolecule #2: X-ray repair cross-complementing protein 5
+Macromolecule #3: DNA-dependent protein kinase catalytic subunit
+Macromolecule #6: DNA repair protein XRCC4
+Macromolecule #7: Non-homologous end-joining factor 1
+Macromolecule #8: Protein PAXX
+Macromolecule #9: DNA ligase 4
+Macromolecule #4: DNA (31-MER)
+Macromolecule #5: DNA (30-MER)
+Macromolecule #10: MAGNESIUM ION
+Macromolecule #11: ADENOSINE-5'-TRIPHOSPHATE
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 2 mg/mL |
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Buffer | pH: 7.9 |
Grid | Model: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: GRAPHENE OXIDE / Support film - topology: CONTINUOUS / Support film - Film thickness: 200 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 10 sec. / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 302 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 100.0 µm / Calibrated defocus max: 4.0 µm / Calibrated defocus min: 2.0 µm / Calibrated magnification: 60000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.0 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 60000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Temperature | Min: 70.0 K / Max: 70.0 K |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 2 / Number real images: 30448 / Average exposure time: 4.0 sec. / Average electron dose: 65.0 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: FLEXIBLE FIT / Target criteria: Cross-correlation coefficient |
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Output model | PDB-8ezb: |