MutS homolog sliding clamps shield the DNA from binding proteins
MutS homolog sliding clamps shield the DNA from binding proteins
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DOI：
10.1074/jbc.ra118.002264
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发表时间：
2018-09-14
影响因子：
4.8
通讯作者：
Fishel, Richard
中科院分区：
文献类型：
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作者：
Hanne, Jeungphill;Britton, Brooke M.;Fishel, Richard
DNA上的滑动夹由进化上保守的酶组成，这些酶协调DNA复制、修复和细胞DNA损伤反应。MutS同源蛋白(MSH)通过识别错配的核苷酸启动错配修复(MMR)，并在ATP存在的情况下形成稳定的滑动夹，沿DNA随机扩散。MSH滑动钳随后加载MutL同源蛋白(MLH/PMS)，形成第二个极其稳定的滑动钳，这些滑动钳共同协调下游MMR成分与距离错配可能数百至数千个核苷酸的切割起始位点。可以想象，错配和远端切除起始点之间的其他蛋白质与DNA的特异性或非特异性结合可能会阻碍这些MMR滑动夹的自由扩散，抑制它们启动修复的能力。在这里，我们使用整体生化分析、单分子荧光成像和数学模型来确定滑动夹子如何克服DNA上的这些障碍。使用细菌和人的MSH蛋白，我们发现增加DNA上的MSH滑动夹的数量减少了大肠杆菌转录抑制因子LacI与其同源启动子LACO的联系。我们的结果提出了一种简单的机制，即MSH滑动夹沿DNA的热扩散改变了其他DNA结合蛋白的结合动力学。这些观察结果似乎普遍适用于DNA上形成的任何稳定的滑动夹。
Sliding clamps on DNA consist of evolutionarily conserved enzymes that coordinate DNA replication, repair, and the cellular DNA damage response. MutS homolog (MSH) proteins initiate mismatch repair (MMR) by recognizing mispaired nucleotides and in the presence of ATP form stable sliding clamps that randomly diffuse along the DNA. The MSH sliding clamps subsequently load MutL homolog (MLH/PMS) proteins that form a second extremely stable sliding clamp, which together coordinate downstream MMR components with the excision-initiation site that may be hundreds to thousands of nucleotides distant from the mismatch. Specific or nonspecific binding of other proteins to the DNA between the mismatch and the distant excision-initiation site could conceivably obstruct the free diffusion of these MMR sliding clamps, inhibiting their ability to initiate repair. Here, we employed bulk biochemical analysis, single-molecule fluorescence imaging, and mathematical modeling to determine how sliding clamps might overcome such hindrances along the DNA. Using both bacterial and human MSH proteins, we found that increasing the number of MSH sliding clamps on a DNA decreased the association of the Escherichia coli transcriptional repressor LacI to its cognate promoter LacO. Our results suggest a simple mechanism whereby thermal diffusion of MSH sliding clamps along the DNA alters the association kinetics of other DNA-binding proteins over extended distances. These observations appear generally applicable to any stable sliding clamp that forms on DNA.