Crystal Structure of DNA-PKcs Reveals a Large Open-Ring Cradle Comprised of HEAT Repeats

Broken chromosomes arising from DNA double strand breaks result from endogenous events such as the production of reactive oxygen species during cellular metabolism, as well as from exogenous sources such as ionizing radiation1, 2, 3. Left unrepaired or incorrectly repaired they can lead to genomic changes that may result in cell death or cancer. DNA-dependent protein kinase (DNA-PK), a holo-enzyme that comprises DNA-dependent protein kinase catalytic subunit (DNA-PKcs)4, 5 and the heterodimer Ku70/Ku80, plays a major role in non-homologous end joining (NHEJ), the main pathway in mammals used to repair double strand breaks6, 7, 8. DNA-PKcs is a serine/threonine protein kinase comprising a single polypeptide chain of 4128 amino acids and belonging to the phosphotidyl inositol 3-kinase (PI3-K)- related protein family9. DNA-PKcs is involved in the sensing and transmission of DNA damage signals to proteins such as p53, setting off events that lead to cell cycle arrest10, 11. It phosphorylates a wide range of substrates in vitro, including Ku70/Ku80, which is translocated along DNA12. Here we present the crystal structure of human DNA-PKcs at 6.6Å resolution, in which the overall fold is for the first time clearly visible. The many α-helical HEAT repeats (helix-turn-helix motifs) facilitate bending and allow the polypeptide chain to fold into a hollow circular structure. The C-terminal kinase domain is located on top of this structure and a small HEAT repeat domain that likely binds DNA is inside. The structure provides a flexible cradle to promote DNA double-strand-break repair.