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Open data
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Basic information
Entry | Database: PDB / ID: 5y3r | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Title | Cryo-EM structure of Human DNA-PK Holoenzyme | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
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![]() | DNA BINDING PROTEIN / Cryo-EM structure / DNA-PK / DNAPKcs / activation / NHEJ | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
Function / homology | ![]() positive regulation of platelet formation / Ku70:Ku80 complex / negative regulation of t-circle formation / T cell receptor V(D)J recombination / DNA end binding / pro-B cell differentiation / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase complex / DNA-dependent protein kinase-DNA ligase 4 complex ...positive regulation of platelet formation / Ku70:Ku80 complex / negative regulation of t-circle formation / T cell receptor V(D)J recombination / DNA end binding / pro-B cell differentiation / small-subunit processome assembly / positive regulation of lymphocyte differentiation / DNA-dependent protein kinase complex / DNA-dependent protein kinase-DNA ligase 4 complex / nonhomologous end joining complex / immunoglobulin V(D)J recombination / immature B cell differentiation / regulation of smooth muscle cell proliferation / cellular response to X-ray / nuclear telomere cap complex / double-strand break repair via alternative nonhomologous end joining / double-strand break repair via classical nonhomologous end joining / regulation of epithelial cell proliferation / telomere capping / Cytosolic sensors of pathogen-associated DNA / IRF3-mediated induction of type I IFN / positive regulation of neurogenesis / regulation of hematopoietic stem cell differentiation / regulation of telomere maintenance / recombinational repair / U3 snoRNA binding / protein localization to chromosome, telomeric region / cellular hyperosmotic salinity response / maturation of 5.8S rRNA / T cell lineage commitment / negative regulation of cGAS/STING signaling pathway / positive regulation of double-strand break repair via nonhomologous end joining / telomeric DNA binding / 2-LTR circle formation / B cell lineage commitment / hematopoietic stem cell proliferation / negative regulation of protein phosphorylation / positive regulation of protein kinase activity / site of DNA damage / Lyases; Carbon-oxygen lyases; Other carbon-oxygen lyases / peptidyl-threonine phosphorylation / 5'-deoxyribose-5-phosphate lyase activity / hematopoietic stem cell differentiation / ATP-dependent activity, acting on DNA / ectopic germ cell programmed cell death / telomere maintenance via telomerase / somitogenesis / mitotic G1 DNA damage checkpoint signaling / neurogenesis / telomere maintenance / DNA helicase activity / activation of innate immune response / cyclin binding / positive regulation of erythrocyte differentiation / cellular response to leukemia inhibitory factor / positive regulation of translation / response to gamma radiation / DNA-dependent protein kinase activity / histone H2AXS139 kinase activity / Nonhomologous End-Joining (NHEJ) / small-subunit processome / enzyme activator activity / cellular response to gamma radiation / regulation of circadian rhythm / protein destabilization / protein-DNA complex / brain development / protein modification process / Hydrolases; Acting on acid anhydrides; Acting on acid anhydrides to facilitate cellular and subcellular movement / double-strand break repair via nonhomologous end joining / cellular response to insulin stimulus / intrinsic apoptotic signaling pathway in response to DNA damage / peptidyl-serine phosphorylation / rhythmic process / double-strand break repair / T cell differentiation in thymus / E3 ubiquitin ligases ubiquitinate target proteins / heart development / double-stranded DNA binding / scaffold protein binding / secretory granule lumen / DNA recombination / transcription regulator complex / RNA polymerase II-specific DNA-binding transcription factor binding / ficolin-1-rich granule lumen / damaged DNA binding / chromosome, telomeric region / transcription cis-regulatory region binding / non-specific serine/threonine protein kinase / protein kinase activity / protein phosphorylation / positive regulation of apoptotic process / ribonucleoprotein complex / protein domain specific binding / innate immune response / protein serine kinase activity / negative regulation of DNA-templated transcription / protein serine/threonine kinase activity / ubiquitin protein ligase binding Similarity search - Function | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
Biological species | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 6.6 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
![]() | Yin, X. / Liu, M. / Tian, Y. / Wang, J. / Xu, Y. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Cryo-EM structure of human DNA-PK holoenzyme. Authors: Xiaotong Yin / Mengjie Liu / Yuan Tian / Jiawei Wang / Yanhui Xu / ![]() Abstract: DNA-dependent protein kinase (DNA-PK) is a serine/threonine protein kinase complex composed of a catalytic subunit (DNA-PKcs) and KU70/80 heterodimer bound to DNA. DNA-PK holoenzyme plays a critical ...DNA-dependent protein kinase (DNA-PK) is a serine/threonine protein kinase complex composed of a catalytic subunit (DNA-PKcs) and KU70/80 heterodimer bound to DNA. DNA-PK holoenzyme plays a critical role in non-homologous end joining (NHEJ), the major DNA repair pathway. Here, we determined cryo-electron microscopy structure of human DNA-PK holoenzyme at 6.6 Å resolution. In the complex structure, DNA-PKcs, KU70, KU80 and DNA duplex form a 650-kDa heterotetramer with 1:1:1:1 stoichiometry. The N-terminal α-solenoid (∼2 800 residues) of DNA-PKcs adopts a double-ring fold and connects the catalytic core domain of DNA-PKcs and KU70/80-DNA. DNA-PKcs and KU70/80 together form a DNA-binding tunnel, which cradles ∼30-bp DNA and prevents sliding inward of DNA-PKcs along with DNA duplex, suggesting a mechanism by which the broken DNA end is protected from unnecessary processing. Structural and biochemical analyses indicate that KU70/80 and DNA coordinately induce conformational changes of DNA-PKcs and allosterically stimulate its kinase activity. We propose a model for activation of DNA-PKcs in which allosteric signals are generated upon DNA-PK holoenzyme formation and transmitted to the kinase domain through N-terminal HEAT repeats and FAT domain of DNA-PKcs. Our studies suggest a mechanism for recognition and protection of broken DNA ends and provide a structural basis for understanding the activation of DNA-PKcs and DNA-PK-mediated NHEJ pathway. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||
History |
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Structure visualization
Movie |
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Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 903.6 KB | Display | ![]() |
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PDB format | ![]() | 715.1 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 847.1 KB | Display | ![]() |
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Full document | ![]() | 1.4 MB | Display | |
Data in XML | ![]() | 211.5 KB | Display | |
Data in CIF | ![]() | 313.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6803MC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
-X-ray repair cross-complementing protein ... , 2 types, 2 molecules AB
#1: Protein | Mass: 57619.352 Da / Num. of mol.: 1 / Fragment: UNP residues 34-534 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() 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: 60972.969 Da / Num. of mol.: 1 / Fragment: UNP residues 6-541 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() 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: 10502.785 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
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#5: DNA chain | Mass: 11042.164 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
-Protein/peptide / Protein , 2 types, 2 molecules KC
#3: Protein/peptide | Mass: 1084.182 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() |
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#6: Protein | Mass: 468885.344 Da / Num. of mol.: 1 / Fragment: UNP residues 10-4128 Source method: isolated from a genetically manipulated source Source: (gene. exp.) ![]() ![]() References: UniProt: P78527, non-specific serine/threonine protein kinase |
-Details
Has protein modification | 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 |
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Sample preparation
Component | Name: PRKDC-Helix / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Source (natural) | Organism: ![]() |
Source (recombinant) | Organism: ![]() |
Buffer solution | pH: 7.4 |
Specimen | Conc.: 0.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 283 K |
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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: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 18000 X / Nominal defocus max: 4700 nm / Nominal defocus min: 1700 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 8 sec. / Electron dose: 50 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of real images: 3496 |
Image scans | Movie frames/image: 32 / Used frames/image: 1-32 |
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Processing
Software | Name: PHENIX / Version: 1.10.1_2155 / Classification: refinement | ||||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: NONE | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 6.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 53451 / Num. of class averages: 1 / Symmetry type: POINT | ||||||||||||||||||||||||||||
Refine LS restraints |
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