Optimizing Oleaginous Yeast Cell Factories for Flavonoids and Hydroxylated Flavonoids Biosynthesis.,ACS Synthetic Biology

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Optimizing Oleaginous Yeast Cell Factories for Flavonoids and Hydroxylated Flavonoids Biosynthesis.
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2019-11-04 , DOI: 10.1021/acssynbio.9b00193
Yongkun Lv _{1,

2,

3} , Monireh Marsafari ₁ , Mattheos Koffas ₄ , Jingwen Zhou _{2,

3} , Peng Xu ₁

Affiliation

Plants possess myriads of secondary metabolites with a broad spectrum of health-promoting benefits. To date, plant extraction is still the primary route to produce high-value natural products which inherently suffers from economics and scalability issues. Heterologous expression of plant biosynthetic gene clusters in microbial host is considered as a feasible approach to overcoming these limitations. Oleaginous yeast produces a large amount of lipid bodies, the abundant membrane structure and the lipophilic environment provide the ideal environment for the regioselectivity and stereoselectivity of many plant-derived P450 enzymes. In this work, we used modular method to construct, characterize, and optimize the flavonoid pathways in Yarrowia lipolytica. We also evaluated various precursor biosynthetic routes and unleashed the metabolic potential of Y. lipolytica to produce flavonoids and hydroxylated flavonoids. Specifically, we have identified that chalcone synthase (CHS) and cytochrome P450 reductases (CPR) were the bottlenecks of hydroxylated flavonoid production. We determined the optimal gene copy number of CHS and CPR to be 5 and 2, respectively. We further removed precursor pathway limitations by expressing genes associated with chorismate and malonyl-CoA supply. With pH and carbon–nitrogen ratio (C/N) optimization, our engineered strain produced 252.4 mg/L naringenin, 134.2 mg/L eriodictyol, and 110.5 mg/L taxifolin from glucose in shake flasks. Flavonoid and its hydroxylated derivatives are most prominently known as antioxidant and antiaging agents. These findings demonstrate our ability to harness the oleaginous yeast as the microbial workhorse to expand nature’s biosynthetic potential, enabling us to bridge the gap between drug discovery and natural product manufacturing.

中文翻译：

优化用于类黄酮和羟基化类黄酮生物合成的含油酵母细胞工厂。

植物具有无数的次级代谢产物，具有广泛的健康促进益处。迄今为止，植物提取仍是生产高价值天然产物的主要途径，而天然产物固有地遭受经济和可扩展性问题的困扰。微生物宿主中植物生物合成基因簇的异源表达被认为是克服这些局限性的可行方法。产油酵母产生大量脂质体，丰富的膜结构和亲脂性环境为许多植物来源的P450酶的区域选择性和立体选择性提供了理想的环境。在这项工作中，我们使用模块化方法来构建，表征和优化解脂耶氏酵母中的类黄酮途径。。我们还评估了各种前体的生物合成途径，并释放了解脂耶氏酵母的代谢潜力。产生类黄酮和羟基化类黄酮。具体来说，我们已经确定查尔酮合酶（CHS）和细胞色素P450还原酶（CPR）是羟化类黄酮产生的瓶颈。我们确定CHS和CPR的最佳基因拷贝数分别为5和2。我们通过表达与分支酸和丙二酰辅酶A供应有关的基因，进一步消除了前体途径的限制。通过优化pH和碳氮比（C / N），我们的工程菌株从摇瓶中的葡萄糖中产生了252.4 mg / L柚皮苷，134.2 mg / L香菊醇和110.5 mg / L滑石粉。类黄酮及其羟基化衍生物最著名的是抗氧化剂和抗衰老剂。这些发现表明，我们有能力利用油质酵母作为微生物的功劳，以扩大自然界的生物合成潜力，

更新日期：2019-11-04

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