Competitive and noncompetitive inhibition of the DNA-dependent protein kinase

Cancer Res. 1999 Jun 1;59(11):2581-6.


The DNA-dependent protein kinase (DNA-PK) is a serine/threonine protein kinase that is involved in mammalian DNA double-strand break repair. The catalytic subunit of DNA-PK (DNA-PKcs) shares sequence homology in its kinase domain with phosphatidylinositol (PI) 3-kinase. Here, we provide a detailed kinetic analysis of DNA-PK inhibition by the PI 3-kinase inhibitor wortmannin and demonstrate this inhibition to be of a noncompetitive nature, with a Ki of 120 nM. Another inhibitor of PI 3-kinase. LY294002, its parent compound, quercetin, and other derivatives have also been studied. These chemicals are competitive inhibitors of DNA-PK, with LY294002 having a Ki of 6.0 microM. Using an antibody to wortmannin, we found that this compound binds covalently to the kinase domain of DNA-PKcs both in vitro and in vivo. Binding of wortmannin to the active site of DNA-PKcs is inhibited by ATP but not by a peptide substrate. Furthermore, wortmannin is able to bind to DNA-PKcs independently of Ku, and it is not stimulated by the presence of DNA. This suggests that the ATP binding site of DNA-PKcs is open constitutively and that DNA activation of the kinase is mediated via another mechanism.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Androstadienes / metabolism
  • Androstadienes / pharmacology*
  • Binding, Competitive
  • Chromones / metabolism
  • Chromones / pharmacology*
  • DNA-Activated Protein Kinase
  • DNA-Binding Proteins*
  • Enzyme Inhibitors / metabolism
  • Enzyme Inhibitors / pharmacology*
  • HL-60 Cells
  • Humans
  • Morpholines / metabolism
  • Morpholines / pharmacology*
  • Nuclear Proteins
  • Protein Serine-Threonine Kinases / antagonists & inhibitors*
  • Protein Serine-Threonine Kinases / metabolism
  • Quercetin / analogs & derivatives
  • Quercetin / pharmacology
  • Radiation-Sensitizing Agents / metabolism
  • Radiation-Sensitizing Agents / pharmacology*
  • Rutin / pharmacology
  • Wortmannin


  • Androstadienes
  • Chromones
  • DNA-Binding Proteins
  • Enzyme Inhibitors
  • Morpholines
  • Nuclear Proteins
  • Radiation-Sensitizing Agents
  • quercitrin
  • 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
  • Rutin
  • Quercetin
  • DNA-Activated Protein Kinase
  • PRKDC protein, human
  • Protein Serine-Threonine Kinases
  • Wortmannin