To mimic the delicately regulated metabolism in nature for improved efficiency, artificial and customized regulatory components for dynamically controlling metabolic networks in multiple layers are essential in laboratory engineering. For this purpose, a novel regulatory component for controlling vanillin biosynthetic pathway was developed through directed evolution, which was responsive to both the product vanillin and substrate ferulic acid, with different capacities. This regulatory component facilitated pathway expression via dynamic control of the intracellular substrate and product concentrations. As vanillin is an antimicrobial compound, low pathway expression and vanillin formation levels enabled better cell growth at an early stage, and the product feedback-activated pathway expression at later stages significantly improved biosynthesis efficiency. This novel multiple-layer dynamic control was demonstrated effective in managing the trade-off between cell growth and production, leading to improved cell growth and vanillin production compared to the conventional or quorum-sensing promoter-controlled pathway. The multiple-layer dynamic control enabled by designed regulatory components responsive to multiple signals shows potential for wide applications in addition to the dynamic controls based on biosynthetic intermediate sensing and quorum sensing reported to date.

为了模仿自然界中微妙的调节代谢以提高效率，在实验室工程中，用于动态控制多层代谢网络的人工和定制调节组件必不可少。为此，通过定向进化开发了用于控制香兰素生物合成途径的新型调节成分，其对产物香兰素和底物阿魏酸都有不同的反应能力。该调节成分通过动态控制细胞内底物和产物浓度来促进途径表达。由于香兰素是一种抗菌化合物，因此低通路表达和香兰素形成水平可以使早期细胞更好地生长，后期阶段的产物反馈激活途径表达显着提高了生物合成效率。与常规的或群体感应的启动子控制的途径相比，这种新颖的多层动态控制被证明可以有效地控制细胞生长与生产之间的折衷，从而改善了细胞生长和香兰素的生产。除了迄今报道的基于生物合成中间感应和群体感应的动态控制外，通过设计的调节组件响应多种信号而实现的多层动态控制还显示出了广泛的应用潜力。与传统的或群体感应的启动子控制的途径相比，导致细胞生长和香兰素生产的改善。除了迄今报道的基于生物合成中间感应和群体感应的动态控制外，通过设计的调节组件响应多种信号而实现的多层动态控制还显示出了广泛的应用潜力。与传统的或群体感应的启动子控制的途径相比，导致细胞生长和香兰素生产的改善。除了迄今报道的基于生物合成中间感应和群体感应的动态控制外，通过设计的调节组件响应多种信号而实现的多层动态控制还显示出了广泛的应用潜力。