Engineering DNA recognition and allosteric response properties of TetR family proteins by using a module-swapping strategy
Engineering DNA recognition and allosteric response properties of TetR family proteins by using a module-swapping strategy
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DOI：
10.1093/nar/gkz666
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发表时间：
2019-09-19
影响因子：
14.9
通讯作者：
Chan, Clement T. Y.
中科院分区：
文献类型：
--
作者：
Dimas, Rey P.;Jordan, Benjamin R.;Chan, Clement T. Y.
合成生物系统的发展需要模块化的生物分子组件来灵活地改变反应途径。在以前的研究中，我们已经建立了一个模块交换的设计原则，工程变构反应和DNA识别性能的调节器之间的LacI家族，其中工程调节器作为实施新的细胞行为的有效组成部分。在这里，我们将这种蛋白质工程化策略引入TetR家族中的两个调节子：TetR（UniProt登录号：P04483）和MphR（Q9 EVJ 6）。TetR DNA结合模块和MphR配体结合模块用于产生TetR-MphR。该混合调节剂具有TetR的DNA结合特性和MphR的配体响应特性，其能够响应细胞中的分子信号来控制基因表达。此外，我们研究了TetR DNA结合模块和MphR配体结合模块之间的分子相互作用，通过使用突变体分析。总之，我们证明了TetR家族调节剂包含离散和功能模块，可用于构建具有新特性的生物组件。这项工作突出了合理设计的效用，作为一种手段，创造模块化的部分细胞工程，并介绍了新的可能性，重新布线细胞反应途径。
The development of synthetic biological systems requires modular biomolecular components to flexibly alter response pathways. In previous studies, we have established a module-swapping design principle to engineer allosteric response and DNA recognition properties among regulators in the LacI family, in which the engineered regulators served as effective components for implementing new cellular behavior. Here we introduced this protein engineering strategy to two regulators in the TetR family: TetR (UniProt Accession ID: P04483) and MphR (Q9EVJ6). The TetR DNA-binding module and the MphR ligand-binding module were used to create the TetR-MphR. This resulting hybrid regulator possesses DNA-binding properties of TetR and ligand response properties of MphR, which is able to control gene expression in response to a molecular signal in cells. Furthermore, we studied molecular interactions between the TetR DNA-binding module and MphR ligand-binding module by using mutant analysis. Together, we demonstrated that TetR family regulators contain discrete and functional modules that can be used to build biological components with novel properties. This work highlights the utility of rational design as a means of creating modular parts for cell engineering and introduces new possibilities in rewiring cellular response pathways.