Orsi Enrico, Beekwilder Jules, van Gelder Dewi, van Houwelingen Adèle, Eggink Gerrit, Kengen Servé W M, Weusthuis Ruud A
Bioprocess Engineering, Wageningen University, 6708PB, Wageningen, The Netherlands.
Wageningen Plant Research, 6700AA, Wageningen, The Netherlands.
Microb Biotechnol. 2020 Jul;13(4):1082-1093. doi: 10.1111/1751-7915.13562. Epub 2020 Mar 24.
Advances in synthetic biology and metabolic engineering have proven the potential of introducing metabolic by-passes within cell factories. These pathways can provide a more efficient alternative to endogenous counterparts due to their insensitivity to host's regulatory mechanisms. In this work, we replaced the endogenous essential 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway for isoprenoid biosynthesis in the industrially relevant bacterium Rhodobacter sphaeroides by an orthogonal metabolic route. The native 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway was successfully replaced by a heterologous mevalonate (MVA) pathway from a related bacterium. The functional replacement was confirmed by analysis of the reporter molecule amorpha-4,11-diene after cultivation with [4- C]glucose. The engineered R. sphaeroides strain relying exclusively on the MVA pathway was completely functional in conditions for sesquiterpene production and, upon increased expression of the MVA enzymes, it reached even higher sesquiterpene yields than the control strain coexpressing both MEP and MVA modules. This work represents an example where substitution of an essential biochemical pathway by an alternative, heterologous pathway leads to enhanced biosynthetic performance.
合成生物学和代谢工程学的进展已证明在细胞工厂中引入代谢旁路的潜力。由于这些途径对宿主调节机制不敏感，它们可以为内源性对应途径提供更有效的替代方案。在这项工作中，我们通过一条正交代谢途径，取代了工业相关细菌球形红杆菌中用于类异戊二烯生物合成的内源性必需2-C-甲基-D-赤藓糖醇4-磷酸（MEP）途径。天然的2-C-甲基-D-赤藓糖醇4-磷酸（MEP）途径成功地被来自相关细菌的异源甲羟戊酸（MVA）途径所取代。在用[4-C]葡萄糖培养后，通过对报告分子amorpha-4,11-diene的分析证实了功能替代。完全依赖MVA途径的工程化球形红杆菌菌株在倍半萜生产条件下完全发挥功能，并且在MVA酶表达增加时，其倍半萜产量甚至比同时共表达MEP和MVA模块的对照菌株更高。这项工作代表了一个例子，即用替代的异源途径取代必需的生化途径可提高生物合成性能。
