Xiao Fengxu, Zhang Yupeng, Zhang Liang, Ding Zhongyang, Shi Guiyang, Li Youran
Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, 214122, People's Republic of China.
National Engineering Research Center for Cereal Fermentation and Food Biomanufacturing, Jiangnan University, 1800 Lihu Avenue, Wuxi, 214122, Jiangsu, People's Republic of China.
Bioresour Bioprocess. 2023 Jan 30;10(1):9. doi: 10.1186/s40643-023-00634-7.
Synthetic biology has rapidly advanced from the setup of native genetic devices to the design of artificial elements able to provide organisms with highly controllable functions. In particular, genetic switches are crucial for deploying new layers of regulation into the engineered organisms. While the assembly and mutagenesis of native elements have been extensively studied, limited progress has been made in rational design of genetic switches due to a lack of understanding of the molecular mechanism by which a specific transcription factor interacts with its target gene. Here, a reliable workflow is presented for designing two categories of genetic elements, one is the switch element-MtlR box and the other is the transcriptional regulatory element- catabolite control protein A (CcpA) box. The MtlR box was designed for ON/OFF-state selection and is controlled by mannitol. The rational design of MtlR box-based molecular structures can flexibly tuned the selection of both ON and OFF states with different output switchability in response to varied kind effectors. Different types of CcpA boxes made the switches with more markedly inducer sensitivities. Ultimately, the OFF-state value was reduced by 90.69%, and the maximum change range in the presence of two boxes was 15.31-fold. This study presents a specific design of the switch, in a plug-and-play manner, which has great potential for controlling the flow of the metabolic pathway in synthetic biology.
合成生物学已从天然遗传装置的构建迅速发展到能够为生物体提供高度可控功能的人工元件的设计。特别是，基因开关对于在工程生物体中部署新的调控层至关重要。虽然对天然元件的组装和诱变已进行了广泛研究，但由于缺乏对特定转录因子与其靶基因相互作用分子机制的了解，基因开关的合理设计进展有限。在此，提出了一种可靠的工作流程，用于设计两类遗传元件，一类是开关元件——MtlR盒，另一类是转录调控元件——分解代谢物控制蛋白A（CcpA）盒。MtlR盒用于开/关状态选择，受甘露醇控制。基于MtlR盒的分子结构的合理设计可以灵活调整开和关状态的选择，以响应不同类型的效应物，具有不同的输出可切换性。不同类型的CcpA盒使开关具有更明显的诱导剂敏感性。最终，关闭状态值降低了90.69%，在存在两个盒的情况下最大变化范围为15.31倍。本研究以即插即用的方式展示了开关的特定设计，在合成生物学中对控制代谢途径的流动具有巨大潜力。
