Climate Change Cluster, University of Technology, 15 Broadway, Ultimo, NSW 2007, Australia.
CSIRO Synthetic Biology Future Science Platform, GPO Box 2583, Brisbane, QLD 4001, Australia.
ACS Synth Biol. 2020 Mar 20;9(3):598-612. doi: 10.1021/acssynbio.9b00455. Epub 2020 Feb 19.
Geraniol is a commercially relevant plant-derived monoterpenoid that is a main component of rose essential oil and used as insect repellent. Geraniol is also a key intermediate compound in the biosynthesis of the monoterpenoid indole alkaloids (MIAs), a group of over 2000 compounds that include high-value pharmaceuticals. As plants naturally produce extremely small amounts of these molecules and their chemical synthesis is complex, industrially sourcing these compounds is costly and inefficient. Hence, microbial hosts suitable to produce MIA precursors through synthetic biology and metabolic engineering are currently being sought. Here, we evaluated the suitability of a eukaryotic microalga, the marine diatom , for the heterologous production of monoterpenoids. Profiling of endogenous metabolism revealed that , unlike other microbes employed for industrial production of terpenoids, accumulates free pools of the precursor geranyl diphosphate. To evaluate the potential for larger synthetic biology applications, we engineered through extrachromosomal, episome-based expression, for the heterologous biosynthesis of the MIA intermediate geraniol. By profiling the production of geraniol resulting from various genetic and cultivation arrangements, reached the maximum geraniol titer of 0.309 mg/L in phototrophic conditions. This work provides (i) a detailed analysis of endogenous terpenoid metabolism, (ii) a successful demonstration of extrachromosomal expression for metabolic pathway engineering with potential gene-stacking applications, and (iii) a convincing proof-of-concept of the suitability of as a novel production platform for heterologous monoterpenoids, with potential for complex pathway engineering aimed at the heterologous production of MIAs.
香叶醇是一种具有商业意义的植物衍生单萜，是玫瑰精油的主要成分，可用作驱虫剂。香叶醇也是生物合成单萜吲哚生物碱（MIAs）的关键中间化合物，MIAs 是一个由 2000 多种化合物组成的群体，其中包括高价值的药物。由于植物自然产生这些分子的量非常少，而且它们的化学合成很复杂，因此工业上获取这些化合物的成本高且效率低。因此，目前正在寻找适合通过合成生物学和代谢工程生产 MIA 前体的微生物宿主。在这里，我们评估了真核微藻海洋硅藻作为异源生产单萜的适宜性。内源性代谢物分析表明，与其他用于萜类化合物工业生产的微生物不同， 积累了前体香叶基二磷酸的游离池。为了评估更大规模的合成生物学应用的潜力，我们通过染色体外、基于附加体的表达工程，使 能够异源生物合成 MIA 中间体香叶醇。通过分析各种遗传和培养安排产生的香叶醇产量， 在光养条件下达到了 0.309mg/L 的最大香叶醇滴度。这项工作提供了 (i) 对 的内源性萜类代谢物的详细分析，(ii) 成功展示了用于代谢途径工程的染色体外表达，具有潜在的基因堆叠应用，以及 (iii) 证明了 作为异源单萜的新型生产平台的适用性，具有针对 MIAs 的异源生产进行复杂途径工程的潜力。
