Long-range communication of Type III restriction enzymes between their cleavage sites based on ATP-triggered 1D diffusion

基于 ATP 触发的一维扩散，III 型限制性内切酶在其切割位点之间进行长距离通讯

• 批准号: 213108456
• 负责人: Professor Dr. Ralf Seidel
• 金额: --
• 依托单位: Peter-Debye-Institut für Physik der weichen Materie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2012
• 资助国家: 德国
• 起止时间: 2011-12-31 至 2014-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/213108456?language=en
• 关键词: Long; range; communication; Type; III

Restriction enzymes are the main defence mechanism of bacteria against invading viruses. They recog-nize viral DNA upon the methylation state of their target sequence and destroy it by cleaving it into pieces. Among the known restriction systems, the Type I and III restriction enzymes use both ATP hy-drolysis in order to carry out DNA cleavage. For Type I restriction enzymes it is meanwhile very well es-tablished that they use ATP hydrolysis to actively translocate DNA, i.e. to act as molecular motors. Whereas a similar mode of action is also proposed for Type III systems, DNA translocation has not been demonstrated so far. Based on indirect measurements we have recently proposed an alternative model in which these enzymes use 1D diffusion rather than translocation to randomly scan large distances on DNA. By combining two single-molecule techniques, in particular magnetic tweezers and fluorescence imaging, we now can directly visualize the diffusion of these enzymes on DNA. This enables us to ad-dress major questions about the mechanisms of these enzymes, such as the role of ATP hydrolysis, con-formational changes that trigger the sliding process as well as the direct communication between en-zymes originating from different target site to collectively cut DNA.

限制性内切酶是细菌抵御入侵病毒的主要防御机制。它们使病毒DNA在其靶序列的甲基化状态下结合，并通过将其切割成碎片来破坏它。在已知的限制系统中，I型和III型限制酶使用ATP水解以进行DNA切割。对于I型限制性内切酶，同时已经很好地确定它们利用ATP水解来主动地易位DNA，即充当分子马达。尽管III型系统也提出了类似的作用模式，但DNA易位迄今尚未得到证实。基于间接测量，我们最近提出了一种替代模型，其中这些酶使用一维扩散而不是易位来随机扫描DNA上的大距离。通过结合两种单分子技术，特别是磁镊和荧光成像，我们现在可以直接观察这些酶在DNA上的扩散。这使我们能够解决关于这些酶的机制的主要问题，例如ATP水解的作用，引发滑动过程的构象变化以及来自不同靶位点的酶之间的直接通信以集体切割DNA。