Three-dimensional structure of the human DNA-PKcs/Ku70/Ku80 complex assembled on DNA and its implications for DNA DSB repair.
DNA-PKcs is a large (approximately 470 kDa) kinase that plays an essential role in the repair of DNA double-strand breaks (DSBs) by nonhomologous end joining (NHEJ). DNA-PKcs is recruited to DSBs by the Ku70/Ku80 heterodimer, with which it forms the core of a multiprotein complex that promotes synapsis of the broken DNA ends. We have purified the human DNA-PKcs/Ku70/Ku80 holoenzyme assembled on a DNA molecule. Its three-dimensional (3D) structure at approximately 25 Angstroms resolution was determined by single-particle electron microscopy. Binding of Ku and DNA elicits conformational changes in the FAT and FATC domains of DNA-PKcs. Dimeric particles are observed in which two DNA-PKcs/Ku70/Ku80 holoenzymes interact through the N-terminal HEAT repeats. The proximity of the dimer contacts to the likely positions of the DNA ends suggests that these represent synaptic complexes that maintain broken DNA ends in proximity and provide a platform for access of the various enzymes required for end processing and ligation.
Spagnolo
Laura
L
Section of Structural Biology and Cancer Research UK DNA Repair Enzyme Research Group, Institute of Cancer Research, Chester Beatty Laboratories, London.
Rivera-Calzada
Angel
A
Pearl
Laurence H
LH
Llorca
Oscar
O
D034961:Antigens, Nuclear / Q000737:chemistry* / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair / Q000502:physiology*; D051747:DNA-Activated Protein Kinase / Q000737:chemistry* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D000072200:Ku Autoantigen; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008958:Models, Molecular; D009687:Nuclear Proteins / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D004247:DNA; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen