Analysis of the kinetic and equilibrium binding of Ku protein to DNA.
The loading of Ku onto a DNA end in a double-strand DNA break is thought to be one of the first steps in the non-homologous DNA end joining (NHEJ) pathway, giving it an essential role in the maintenance of genomic integrity. The binding of Ku to DNA is complicated since DNA can accommodate multiple Ku subunits, which can translocate on the DNA strand. Furthermore, Ku may exhibit cooperativity in the loading process. Therefore, simple one- to-one kinetic models are unable to adequately simulate the process. However, through the use of computer simulation and curve-fitting, we are able to provide a comprehensive mechanistic model and rate constants that closely approximate experimental data for DNA molecules that bind one, two, and three Ku molecules under both kinetic and equilibrium conditions. The model obtains a best fit with Ku having a roughly seven-fold preference to bind to DNA ends rather than internal positions and is consistent with Ku having a strong preference of which face of the protein loads onto the DNA end.
Taghva
Alexander
A
Department of Pathology, Norris Comprehensive Cancer Center, University of Southern California, Rm. 5428, Los Angeles, CA 90089, USA.
Ma
Yunmei
Y
Lieber
Michael R
MR
D000818:Animals; D034961:Antigens, Nuclear; D003198:Computer Simulation; D004247:DNA / Q000378:metabolism*; D004265:DNA Helicases; D004260:DNA Repair; D004268:DNA-Binding Proteins / Q000378:metabolism*; D000072200:Ku Autoantigen; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000378:metabolism*
D034961:Antigens, Nuclear; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D004247:DNA; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen