Debottlenecking mevalonate pathway for antimalarial drug precursor amorphadiene biosynthesis in Yarrowia lipolytica
Debottlenecking mevalonate pathway for antimalarial drug precursor amorphadiene biosynthesis in Yarrowia lipolytica
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DOI：
10.1016/j.mec.2019.e00121
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发表时间：
2020-06-01
影响因子：
5.2
通讯作者：
Xu, Peng
中科院分区：
文献类型：
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作者：
Marsafari, Monireh;Xu, Peng
世界卫生组织报告说，发展中国家一半的人口面临感染疟疾的危险。青蒿素是最有效的抗疟疾药物，是从植物黄花蒿中提取的倍半萜类内过氧化物。由于可扩展性和经济问题，植物提取或化学合成无法为青蒿素的大规模生产提供可持续的途径。由于地缘政治和气候因素，青蒿素的价格一直在200美元/公斤至1100美元/公斤之间波动。微生物发酵被认为是稳定青蒿素供应链的一种有前途的方法。解脂耶氏酵母（Yarrowia lipolytica）是一种产油酵母，具有产生大量油脂和油脂化学物质的能力。本文对Y.解脂菌用于紫穗槐二烯生产。基因过表达表明HMG-CoA和乙酰CoA供应是紫穗槐二烯生产的两个限制性瓶颈。我们已经确定了最佳的HMG-CoA还原酶，并确定了前体途径的最佳基因拷贝数。通过抑制脂肪酸合酶或激活脂肪酸降解途径，进一步改善了阿莫非二烯的生产。通过共表达甲羟戊酸激酶（由Erg 12编码），推拉策略使工程菌株能够在摇瓶中产生171.5 mg/L的紫穗槐二烯。这些结果表明，平衡碳通量和操纵前体竞争途径是提高产油酵母中紫穗槐二烯生物合成的关键因素;解脂菌是一种很有前途的微生物宿主，可以扩大自然界的生物合成能力，使我们能够快速获得抗疟药物前体。
World Health Organization reports that half of the population in developing countries are at risk of malaria infection. Artemisinin, the most potent anti-malaria drug, is a sesquiterpene endoperoxide extracted from the plant Artemisia annua. Due to scalability and economics issues, plant extraction or chemical synthesis could not provide a sustainable route for large-scale manufacturing of artemisinin. The price of artemisinin has been fluctuating from 200$/Kg to 1100$/Kg, due to geopolitical and climate factors. Microbial fermentation was considered as a promising method to stabilize the artemisinin supply chain. Yarrowia lipolytica, is an oleaginous yeast with proven capacity to produce large quantity of lipids and oleochemicals. In this report, the lipogenic acetyl-CoA pathways and the endogenous mevalonate pathway of Y. lipolytica were harnessed for amorphadiene production. Gene overexpression indicate that HMG-CoA and acetyl-CoA supply are two limiting bottlenecks for amorphadiene production. We have identified the optimal HMG-CoA reductase and determined the optimal gene copy number for the precursor pathways. Amorphadiene production was improved further by either inhibiting fatty acids synthase or activating the fatty acid degradation pathway. With co-expression of mevalonate kinase (encoded by Erg12), a push-and-pull strategy enabled the engineered strain to produce 171.5 mg/L of amorphadiene in shake flasks. These results demonstrate that balancing carbon flux and manipulation of precursor competing pathways are key factors to improve amorphadiene biosynthesis in oleaginous yeast; and Y. lipolytica is a promising microbial host to expand nature's biosynthetic capacity, allowing us to quickly access antimalarial drug precursors.