文献类型：期刊文献

中文题名：毕赤酵母生产二十碳五烯酸细胞工厂的构建与优化

英文题名：Construction and optimization of a microbial cell factory for eicosapentaenoic acid production in Pichia pastoris

作者：黄婕[1];侯淑婷[1];李慧[1];周伟[1];凌睿婧[1];杨意琛[1];高蓓[1]

机构：[1]华东理工大学生物工程学院鲁华生物技术研究所,上海201306

年份：2025

卷号：41

期号：5

起止页码：1908

中文期刊名：生物工程学报

外文期刊名：Chinese Journal of Biotechnology

收录：;EI(收录号：20252218504594);北大核心:【北大核心2023】；

基金：国家重点研发计划(2024YFA0917900);上海自然科学基金(21ZR1416500)。

语种：中文

中文关键词：长链多不饱和脂肪酸;二十碳五烯酸;毕赤酵母;合成生物学;细胞工厂

外文关键词：long-chain polyunsaturated fatty acids;eicosapentaenoic acid;Pichia pastoris;synthetic biology;cell factories

摘要：二十碳五烯酸(eicosapentaenoic acid,EPA)是一种ω-3长链多不饱和脂肪酸,因其多种生理活性而备受关注。EPA的传统来源是从深海鱼油中提取,但随着鱼类资源的枯竭和环境污染的加剧,该方法越来越不可持续。为了寻找可持续的绿色生产方式,本研究构建了毕赤酵母异源合成EPA的细胞工厂,通过敲除脂肪酸下游代谢途径和竞争途径提高了工程菌株的总脂肪酸(total fatty acids,TFAs)的积累,同时强化了C16/18延伸酶、C18/20延伸酶和去饱和酶的表达,工程菌株在以葡萄糖为碳源的条件下积累了677.34 mg/L TFAs、60.81 mg/L EPA。进一步过表达以上3种关键酶,EPA的产量达到68.89 mg/L。同时,使用醇氧化酶启动子PAOX1替换3个关键酶的葡萄糖启动子P_(GAP)探究了甲醇诱导下的EPA的合成水平,EPA的产量达到75.21 mg/L。本研究首次实现了在毕赤酵母中从头合成EPA,为构建利用一碳资源甲醇生产EPA的细胞工厂提供了有益参考。
Eicosapentaenoic acid(EPA)is anω-3 long-chain polyunsaturated fatty acid valued for its diverse physiological activities.The production of EPA by extraction from marine fish oils is becoming increasingly unsustainable due to depleting fish stocks and rising environmental pollution.Consequently,alternative sources for EPA production are necessary.In this study,we constructed a Pichia pastoris cell factory for heterologous production of EPA.To enhance the accumulation of total fatty acids(TFAs)in the engineered strain,we knocked out the downstream metabolic pathway of fatty acids and the competitive pathway.Simultaneously,the expression of C16/18 elongase,C18/20 elongase,and desaturase were strengthened.With glucose as the carbon source,the engineered strain accumulated 677.34 mg/L TFAs and 60.81 mg/L EPA.The three key enzymes were further overexpressed to increase the EPA production,which reached a titer of 68.89 mg/L.Furthermore,we replaced the glucose promoter P_(GAP)of three key enzymes in the EPA synthesis pathway with the promoter PAOX1 of the alcohol oxidase AOX1 to explore the methanol-induced EPA synthesis,which further increased the EPA titer to 75.21 mg/L.This study lays a foundation for the biosynthesis of long-chain polyunsaturated fatty acids in P.pastoris and the construction of P.pastoris cell factories for producing EPA with methanol,a one-carbon resource.