环氧角鲨烯合成通路在大肠杆菌中的构建和优化

Construction and optimization of squalene epoxide synthetic pathway in Escherichia coli

摘要:

三萜类化合物是一类广泛应用于医药、保健和化妆品等行业的天然产物，具有巨大的商业价值。生物合成三萜类化合物依赖于环氧角鲨烯的高效合成。角鲨烯环氧化酶是整个合成途径中的关键酶，其催化NADPH依赖的环氧化反应将角鲨烯转变为环氧角鲨烯。通过筛选不同来源的角鲨烯环氧化酶，截短的大鼠角鲨烯环氧化酶 (RnSETC) 在大肠杆菌Escherichia coli工程菌中表现出最强的活性；进一步考察内源性细胞色素P450还原酶样 (CPRL) 蛋白对环氧角鲨烯合成的影响显示，在中度拷贝质粒上以Lac启动子调控NADH：醌氧化还原酶 (WrbA) 的表达使得环氧角鲨烯产量提高近2.5倍。研究结果表明，所构建的环氧角鲨烯合成途径可以在大肠杆菌中实现三萜类合成关键前体环氧角鲨烯的合成，为生物合成三萜类化合物提供了重要借鉴。

Abstract:

Triterpenoids are a class of natural products of great commercial value that are widely used in pharmaceutical, health care and cosmetic industries. The biosynthesis of triterpenoids relies on the efficient synthesis of squalene epoxide, which is synthesized from the NADPH dependent oxidation of squalene catalyzed by squalene epoxidase. We screened squalene epoxidases derived from different species, and found the truncated squalene epoxidase from Rattus norvegicus (RnSETC) showed the highest activity in engineered Escherichia coli. Further examination of the effect of endogenous cytochrome P450 reductase like (CPRL) proteins showed that overexpression of NADH: quinone oxidoreductase (WrbA) under Lac promoter in a medium-copy number plasmid increased the production of squalene epoxide by nearly 2.5 folds. These results demonstrated that the constructed pathway led to the production of squalene epoxide, an important precursor for the biosynthesis of triterpenoids.