In mammalian cells, double-strand break repair and V(D)J recombination require DNA-dependent protein kinase (DNA-PK), a serine/threonine kinase that is activated by DNA. DNA-PK consists of a 460-kDa subunit (p460) that contains a putative kinase domain and a heterodimeric subunit (Ku) that binds to double-stranded DNA ends. Previous reports suggested that the activation of DNA-PK requires the binding of Ku to DNA. To investigate this further, p460 and Ku were purified separately to homogeneity. Surprisingly, p460 was capable of binding to DNA in the absence of Ku. The binding of p460 to double-stranded DNA ends was salt-labile and could be disrupted by single-stranded or supercoiled DNA, properties distinct from the binding of Ku to DNA. Under low salt conditions, which permitted the binding of p460 to DNA ends, the kinase was activated. Under higher salt conditions, which inhibited the binding of p460, activation of the kinase required the addition of Ku. Significantly, when the length of DNA decreased to 22 bp, Ku competed with p460 for DNA binding and inhibited kinase activity. These data demonstrate that p460 is a self-contained kinase that is activated by direct interaction with double-stranded DNA and that the role of Ku is to stabilize the binding of p460 to DNA ends.
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Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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Site | Modification | Modifier | Source | Reference |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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