Cancer Research UK DNA Repair Enzymes Group, The Institute of Cancer Research, London SW3 6JB, UK.
Nucleic Acids Res. 2012 May;40(9):4168-77. doi: 10.1093/nar/gkr1231. Epub 2012 Jan 5.
The DNA-dependent protein kinase (DNA-PK) and Poly(ADP-ribose) polymerase-1 (PARP1) are critical enzymes that reduce genomic damage caused by DNA lesions. They are both activated by DNA strand breaks generated by physiological and environmental factors, and they have been shown to interact. Here, we report in vivo evidence that DNA-PK and PARP1 are equally necessary for rapid repair. We purified a DNA-PK/PARP1 complex loaded on DNA and performed electron microscopy and single particle analysis on its tetrameric and dimer-of-tetramers forms. By comparison with the DNA-PK holoenzyme and fitting crystallographic structures, we see that the PARP1 density is in close contact with the Ku subunit. Crucially, PARP1 binding elicits substantial conformational changes in the DNA-PK synaptic dimer assembly. Taken together, our data support a functional, in-pathway role for DNA-PK and PARP1 in double-strand break (DSB) repair. We also propose a NHEJ model where protein-protein interactions alter substantially the architecture of DNA-PK dimers at DSBs, to trigger subsequent interactions or enzymatic reactions.
DNA 依赖性蛋白激酶（DNA-PK）和聚（ADP-核糖）聚合酶 1（PARP1）是减少由 DNA 损伤引起的基因组损伤的关键酶。它们都被生理和环境因素产生的 DNA 链断裂所激活，并且已经证明它们相互作用。在这里，我们报告了体内证据，表明 DNA-PK 和 PARP1 对于快速修复同样重要。我们纯化了加载在 DNA 上的 DNA-PK/PARP1 复合物，并对其四聚体和四聚体二聚体形式进行了电子显微镜和单颗粒分析。通过与 DNA-PK 全酶的比较和晶体结构的拟合，我们发现 PARP1 密度与 Ku 亚基密切接触。至关重要的是，PARP1 的结合会引起 DNA-PK 突触二聚体组装的显著构象变化。总之，我们的数据支持 DNA-PK 和 PARP1 在双链断裂（DSB）修复中具有功能上的、通路内的作用。我们还提出了一个 NHEJ 模型，其中蛋白质-蛋白质相互作用会极大地改变 DSB 处 DNA-PK 二聚体的结构，以触发随后的相互作用或酶反应。
