Swammerdam Institute for Life Sciences, University of Amsterdam, Amsterdam, Netherlands.
J Cell Biol. 2010 May 3;189(3):445-63. doi: 10.1083/jcb.200909175.
To understand how multiprotein complexes assemble and function on chromatin, we combined quantitative analysis of the mammalian nucleotide excision DNA repair (NER) machinery in living cells with computational modeling. We found that individual NER components exchange within tens of seconds between the bound state in repair complexes and the diffusive state in the nucleoplasm, whereas their net accumulation at repair sites evolves over several hours. Based on these in vivo data, we developed a predictive kinetic model for the assembly and function of repair complexes. DNA repair is orchestrated by the interplay of reversible protein-binding events and progressive enzymatic modifications of the chromatin substrate. We demonstrate that faithful recognition of DNA lesions is time consuming, whereas subsequently, repair complexes form rapidly through random and reversible assembly of NER proteins. Our kinetic analysis of the NER system reveals a fundamental conflict between specificity and efficiency of chromatin-associated protein machineries and shows how a trade off is negotiated through reversibility of protein binding.
为了了解多蛋白复合物如何在染色质上组装和发挥作用，我们将哺乳动物核苷酸切除 DNA 修复 (NER) 机制的定量分析与计算建模相结合。我们发现，单个 NER 组件在修复复合物的结合态和核质的扩散态之间交换仅需数十秒，而它们在修复部位的净积累则需要数小时。基于这些体内数据，我们开发了一个用于修复复合物组装和功能的预测性动力学模型。DNA 修复是由可逆的蛋白质结合事件和染色质底物的酶促修饰的相互作用来协调的。我们证明，DNA 损伤的准确识别需要耗费时间，而随后，通过 NER 蛋白的随机和可逆组装，修复复合物迅速形成。我们对 NER 系统的动力学分析揭示了染色质相关蛋白机器的特异性和效率之间的基本冲突，并展示了如何通过蛋白质结合的可逆性来协商这种权衡。
