Biotransformation Innovation Platform (BioTrans), Agency for Science, Technology, and Research (A*STAR), Singapore, Singapore.
Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore, Singapore.
Biotechnol Bioeng. 2018 Jan;115(1):174-183. doi: 10.1002/bit.26462.
Apocarotenoids, such as α-, β-ionone, and retinol, have high commercial values in the food and cosmetic industries. The demand for natural ingredients has been increasing dramatically in recent years. However, attempts to overproduce β-ionone in microorganisms have been limited by the complexity of the biosynthetic pathway. Here, an Escherichia coli-based modular system was developed to produce various apocarotenoids. Incorporation of enzyme engineering approaches (N-terminal truncation and protein fusion) into modular metabolic engineering strategy significantly improved α-ionone production from 0.5 mg/L to 30 mg/L in flasks, producing 480 mg/L of α-ionone in fed-batch fermentation. By modifying apocarotenoid genetic module, this platform strain was successfully re-engineered to produce 32 mg/L and 500 mg/L of β-ionone in flask and bioreactor, respectively (>80-fold higher than previously reported). Similarly, 33 mg/L of retinoids was produced in flask by reconstructing apocarotenoid module, demonstrating the versatility of the "plug-n-play" modular system. Collectively, this study highlights the importance of the strategy of simultaneous modular pathway optimization and enzyme engineering to overproduce valuable chemicals in microbes.
类胡萝卜素衍生物，如α-、β-紫罗兰酮和视黄醇，在食品和化妆品行业具有很高的商业价值。近年来，对天然成分的需求急剧增加。然而，由于生物合成途径的复杂性，试图在微生物中过量生产β-紫罗兰酮的尝试受到了限制。在这里，开发了基于大肠杆菌的模块化系统来生产各种类胡萝卜素衍生物。将酶工程方法（N 端截断和蛋白融合）纳入模块化代谢工程策略中，可显著提高α-紫罗兰酮的产量，从摇瓶中的 0.5mg/L 提高到 30mg/L，在分批补料发酵中产生 480mg/L 的α-紫罗兰酮。通过修饰类胡萝卜素遗传模块，该平台菌株被成功重新设计，在摇瓶和生物反应器中分别产生 32mg/L 和 500mg/L 的β-紫罗兰酮（比之前报道的提高了 80 多倍）。同样，通过重构类胡萝卜素模块，在摇瓶中生产了 33mg/L 的视黄醇，证明了“即插即用”模块化系统的多功能性。总的来说，这项研究强调了同时优化模块化途径和酶工程策略以在微生物中过量生产有价值化学品的重要性。
