Wang Lijuan, Deng Yongdong, Peng Rihe, Gao Jianjie, Li Zhenjun, Zhang Wenhui, Xu Jing, Wang Bo, Wang Yu, Han Hongjuan, Fu Xiaoyan, Tian Yongsheng, Yao Quanhong
Shanghai Key Laboratory of Agricultural Genetics and Breeding, Biotechnology Research Institute of Shanghai Academy of Agricultural Sciences, 2901 Beidi Road, Shanghai, China.
Key Laboratory for Safety Assessment (Environment) of Agricultural Genetically Modified Organisms, Ministry of Agriculture and Rural Affairs, Shanghai, China.
Biotechnol Biofuels Bioprod. 2023 Mar 2;16(1):34. doi: 10.1186/s13068-023-02284-5.
Terrequinone A is a bis-indolylquinone natural product with antitumor activity. Due to its unique asymmetric quinone core structure and multiple functional groups, biosynthesis is more efficient and environmentally friendly than traditional chemical synthesis. Currently, most bis-indolylquinones are obtained by direct extraction from fungi or by chemical synthesis. By focusing on the biosynthesis of terrequinone A, we hope to explore the way to synthesize bis-indolylquinones de novo using Escherichia coli as a cell factory.
In this study, a terrequinone A synthesis pathway containing the tdiA-tdiE genes was constructed into Escherichia coli and activated by a phosphopantetheinyl transferase gene sfp, enabling the strain to synthesize 1.54 mg/L of terrequinone A. Subsequently, a two-step isopentenol utilization pathway was introduced to enhance the supply of endogenous dimethylallyl diphosphate (DMAPP) in E. coli, increasing the level of terrequinone A to 20.1 mg/L. By adjusting the L-tryptophan (L-Trp)/prenol ratio, the major product could be changed from ochrindole D to terrequinone A, and the content of terrequinone A reached the highest 106.3 mg/L under the optimized culture conditions. Metabolic analysis of L-Trp indicated that the conversion of large amounts of L-Trp to indole was an important factor preventing the further improvement of terrequinone A yield.
A comprehensive approach was adopted and terrequinone A was successfully synthesized from low-cost L-Trp and prenol in E. coli. This study provides a metabolic engineering strategy for the efficient synthesis of terrequinone A and other similar bis-indolylquinones with asymmetric quinone cores. In addition, this is the first report on the de novo biosyhthesis of terrequinone A in an engineered strain.
特瑞喹酮A是一种具有抗肿瘤活性的双吲哚基醌天然产物。由于其独特的不对称醌核心结构和多个官能团，生物合成比传统化学合成更高效、更环保。目前，大多数双吲哚基醌是通过从真菌中直接提取或化学合成获得的。通过关注特瑞喹酮A的生物合成，我们希望探索以大肠杆菌作为细胞工厂从头合成双吲哚基醌的方法。
在本研究中，将包含tdiA - tdiE基因的特瑞喹酮A合成途径构建到大肠杆菌中，并通过磷酸泛酰巯基乙胺基转移酶基因sfp进行激活，使该菌株能够合成1.54 mg/L的特瑞喹酮A。随后，引入两步异戊烯醇利用途径以增强大肠杆菌内源性二甲基烯丙基二磷酸（DMAPP）的供应，将特瑞喹酮A的水平提高到20.1 mg/L。通过调整L - 色氨酸（L - Trp）/异戊烯醇比例，主要产物可从奥克吲哚D转变为特瑞喹酮A，并且在优化的培养条件下特瑞喹酮A的含量达到最高106.3 mg/L。L - Trp的代谢分析表明，大量L - Trp转化为吲哚是阻碍特瑞喹酮A产量进一步提高的重要因素。
采用了一种综合方法，成功地在大肠杆菌中从低成本的L - Trp和异戊烯醇合成了特瑞喹酮A。本研究为高效合成特瑞喹酮A和其他具有不对称醌核心的类似双吲哚基醌提供了一种代谢工程策略。此外，这是关于工程菌株中特瑞喹酮A从头生物合成的首次报道。
