Systems Metabolic Engineering of Escherichia coli Coculture for De Novo Production of Genistein
Systems Metabolic Engineering of Escherichia coli Coculture for De Novo Production of Genistein
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用于从头生产金雀异黄酮的大肠杆菌共培养系统代谢工程
DOI：
10.1021/acssynbio.1c00590
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发表时间：
2022-05-20
影响因子：
4.7
通讯作者：
Zhao, Guang-Rong
中科院分区：
文献类型：
--
作者：
Liu, Xue;Li, Lingling;Zhao, Guang-Rong
染料木黄酮是一种植物源化合物，具有抗衰老、抗肿瘤、抗神经退行性疾病和抗炎症等多种生物活性。作为一种典型的复杂类黄酮，其从糖的微生物生产仍有待完成。本研究采用系统代谢工程策略，设计并建立了一种三株嗜热大肠杆菌共培养体系，首次实现了染料木素的从头合成。首先，我们通过筛选和整合查耳酮异构酶样蛋白（一种辅助成分，用于纠正查耳酮合酶混杂）来重建柚皮素模块。此外，我们通过对植物P450酶2-羟基异黄烷酮合酶和细胞色素P450酶还原酶的N端修饰，设计并构建了染料木黄酮模块。将柚皮素产生菌与苯丙氨酸营养缺陷型的对香豆酸高产菌共培养，两菌共培养通过单向营养流产生共生现象，有利于以葡萄糖为底物高效生产柚皮素，其效价为206.5mg/L。随后设计了一种三菌株的共生共培养物，该共培养物从葡萄糖和甘油混合物中产生迄今为止最高滴度的60.8 mg/L染料木黄酮。该共培养体系为黄酮类化合物的生产提供了一个灵活多样的平台，为工程菌生产复杂天然产物提供了一个广阔的前景。杆菌
: Genistein is a plant-derived isoflavone possessing various bioactivities to prevent aging, carcinogenesis, and neurodegenerative and inflammation diseases. As a typical complex flavonoid, its microbial production from sugar remains to be completed. Here, we use systems metabolic engineering stategies to design and develop a three-strain commensalistic Escherichia coli coculture that for the first time realized the de novo production of genistein. First, we reconstituted the naringenin module by screening and incorporating chalcone isomerase-like protein, an auxiliary component to rectify the chalcone synthase promiscuity. Furthermore, we devised and constructed the genistein module by N-terminal modifications of plant P450 enzyme 2- hydroxyisoflavanone synthase and cytochrome P450 enzyme reductase. When naringenin-producing strain was cocultivated with pcoumaric acid-overproducing strain (a phenylalanine-auxotroph), two-strain coculture worked as commensalism through a unidirectional nutrient flow, which favored the efficient production of naringenin with a titer of 206.5 mg/L from glucose. A threestrain commensalistic coculture was subsequently engineered, which produced the highest titer to date of 60.8 mg/L genistein from a glucose and glycerol mixture. The commensalistic coculture is a flexible and versatile platform for the production of flavonoids, indicating a promising future for production of complex natural products in engineered E. coli.