Nguyen Thang Minh, Nakata Eiji, Zhang Zhengxiao, Saimura Masayuki, Dinh Huyen, Morii Takashi
Institute of Advanced Energy , Kyoto University , Uji , Kyoto 611-0011 , Japan . Email:
Chem Sci. 2019 Aug 20;10(40):9315-9325. doi: 10.1039/c9sc02990g. eCollection 2019 Oct 28.
Sequence-selective chemical modification of DNA by synthetic ligands has been a long-standing challenge in the field of chemistry. Even when the ligand consists of a sequence-specific DNA binding domain and reactive group, sequence-selective reactions by these ligands are often accompanied by off-target reactions. A basic principle to design DNA modifiers that react at specific sites exclusively governed by DNA sequence recognition remains to be established. We have previously reported selective DNA modification by a self-ligating protein tag conjugated with a DNA-binding domain, termed as a modular adaptor, and orthogonal application of modular adaptors by relying on the chemoselectivity of the protein tag. The sequence-specific crosslinking reaction by the modular adaptor is thought to proceed in two steps: the first step involves the formation of a DNA-protein complex, while in the second step, a proximity-driven intermolecular crosslinking occurs. According to this scheme, the specific crosslinking reaction of a modular adaptor would be driven by the DNA recognition process only when the dissociation rate of the DNA complex is much higher than the rate constant for the alkylation reaction. In this study, as a proof of principle, a set of combinations for modular adaptors and their substrates were utilized to evaluate the reactions. Three types of modular adaptors consisting of a single type of self-ligating tag and three types of DNA binding proteins fulfill the kinetic requirements for the reaction of the self-ligating tag with a substrate and the dissociation of the DNA-protein complex. These modular adaptors actually undergo sequence-specific crosslinking reactions exclusively driven by the recognition of a specific DNA sequence. The design principle of sequence-specific modular adaptors based on the kinetic aspects of complex formation and chemical modification is applicable for developing recognition-driven selective modifiers for proteins and other biological macromolecules.
通过合成配体对DNA进行序列选择性化学修饰一直是化学领域长期存在的挑战。即使配体由序列特异性DNA结合结构域和反应基团组成，这些配体的序列选择性反应也常常伴随着脱靶反应。设计仅在由DNA序列识别专门控制的特定位点发生反应的DNA修饰剂的基本原理仍有待确立。我们之前报道了一种与DNA结合结构域偶联的自连接蛋白标签对DNA的选择性修饰，该标签被称为模块化接头，并且通过依赖蛋白标签的化学选择性实现了模块化接头的正交应用。模块化接头的序列特异性交联反应被认为分两步进行：第一步涉及形成DNA-蛋白质复合物，而在第二步中，发生邻近驱动的分子间交联。根据该方案，仅当DNA复合物的解离速率远高于烷基化反应的速率常数时，模块化接头的特异性交联反应才会由DNA识别过程驱动。在本研究中，作为原理验证，利用了一组模块化接头及其底物的组合来评估反应。由单一类型的自连接标签和三种类型的DNA结合蛋白组成的三种类型的模块化接头满足了自连接标签与底物反应以及DNA-蛋白质复合物解离的动力学要求。这些模块化接头实际上仅通过对特定DNA序列的识别而发生序列特异性交联反应。基于复合物形成和化学修饰的动力学方面的序列特异性模块化接头的设计原理适用于开发用于蛋白质和其他生物大分子的识别驱动的选择性修饰剂。
