Chan D W, Lees-Miller S P
Department of Biological Sciences, University of Calgary, 2500 University Drive, N.W., Calgary, Alberta, T2N 1N4, Canada.
J Biol Chem. 1996 Apr 12;271(15):8936-41. doi: 10.1074/jbc.271.15.8936.
The DNA-dependent protein kinase (DNA-PK) requires for activity free ends or other discontinuities in the structure of double strand DNA. In vitro, DNA-PK phosphorylates several transcription factors and other DNA-binding proteins and is thought to function in DNA damage recognition or repair and/or transcription. Here we show that in vitro DNA-PK undergoes autophosphorylation of all three protein subunits (DNA-PKcs, Ku p70 and Ku p80) and that phosphorylation correlates with inactivation of the serine/threonine kinase activity of DNA-PK. Significantly, activity is restored by the addition of purified native DNA-PKcs but not Ku, suggesting that inactivation is due to autophosphorylation of DNA-PKcs. Our data also suggest that autophosphorylation results in dissociation of DNA-PKcs from the Ku-DNA complex. We suggest that autophosphorylation is an important mechanism for the regulation of DNA-PK activity.
DNA依赖性蛋白激酶（DNA-PK）的活性需要双链DNA结构中的游离末端或其他不连续处。在体外，DNA-PK使几种转录因子和其他DNA结合蛋白磷酸化，并被认为在DNA损伤识别或修复和/或转录中发挥作用。在这里，我们表明在体外DNA-PK会对所有三个蛋白质亚基（DNA-PKcs、Ku p70和Ku p80）进行自身磷酸化，并且这种磷酸化与DNA-PK的丝氨酸/苏氨酸激酶活性的失活相关。重要的是，通过添加纯化的天然DNA-PKcs而非Ku可恢复活性，这表明失活是由于DNA-PKcs的自身磷酸化。我们的数据还表明，自身磷酸化导致DNA-PKcs从Ku-DNA复合物中解离。我们认为自身磷酸化是调节DNA-PK活性的一种重要机制。
