On the mechanisms of Ku protein binding to DNA.
The in vitro DNA-binding activity of Ku protein, a heterodimer of 70 and 86 kDa subunits, was studied using affinity-purified protein. Ku protein bound to different DNA probes and displayed a multiple-band pattern in band mobility shift assays. The protein-DNA complex formation was effectively blocked by different DNA competitors, indicating a non-sequence specific binding of Ku protein to DNA; no preference of binding of Ku protein to regulatory sequences derived from U1 snRNA, U6 snRNA or nucleolar protein p120 genes was observed. The number and size of the Ku protein-DNA complexes increased with increasing of the protein concentration and the size of DNA probe, suggesting that the protein accumulates on the DNA fragment until saturation of the binding sites. In UV-crosslinking experiments, the binding of Ku protein to DNA was shown to start with the 70 kDa subunit contacting free DNA ends.
Zhang
W W
WW
Department of Pharmacology, Baylor College of Medicine, Houston, Texas 77030.
Yaneva
M
M
D034961:Antigens, Nuclear; D001667:Binding, Competitive; D002846:Chromatography, Affinity; D004247:DNA / Q000378:metabolism*; D004265:DNA Helicases; D015342:DNA Probes; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005796:Genes; D006367:HeLa Cells; D006801:Humans; D000072200:Ku Autoantigen; D046911:Macromolecular Substances; D008970:Molecular Weight; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction / Q000379:methods; D011485:Protein Binding; D012342:RNA, Small Nuclear / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D014466:Ultraviolet Rays
D034961:Antigens, Nuclear; D015342:DNA Probes; D004268:DNA-Binding Proteins; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D012342:RNA, Small Nuclear; D004247:DNA; D004265:DNA Helicases; C106893:XRCC5 protein, human; C000606509:Xrcc6 protein, human; D000072200:Ku Autoantigen