代谢工程改造酿酒酵母生产β-胡萝卜素

Production of β-carotene by metabolically engineered Saccharomyces cerevisiae

摘要:

β-胡萝卜素在食品、药品和化妆品领域有广泛用途。为获得生产β-胡萝卜素的微生物细胞工厂，本研究首先在酿酒酵母BY4742中过表达甲羟戊酸 (MVA) 途径的限速酶3-羟基-3-甲基戊二酰辅酶A还原酶 (HMGR) 基因及二萜化合物合成的关键酶牻牛儿基牻牛儿基焦磷酸合酶 (GGPS) 基因，来提高牻牛儿基牻牛儿基焦磷酸 (GGPP) 的供给。在酿酒酵母底盘菌BY4742-T2的基础上整合来源于成团泛菌和红法夫酵母的β-胡萝卜素合成基因，比较酿酒酵母工程菌生产β-胡萝卜素的差别。结果表明提高酿酒酵母中HMGR和GGPS酶基因的表达能将工程菌中β-胡萝卜素的产量提高26.0倍。另外，来源于真核生物红法夫酵母的合成基因相比成团泛菌，更有利于酿酒酵母生产β-胡萝卜素。最终获得的酿酒酵母工程菌BW02能生产1.56 mg/g干细胞重的β-胡萝卜素，为进一步获得高产β-胡萝卜素细胞工厂提供基础。

Abstract:

β-carotene has a wide range of application in food, pharmaceutical and cosmetic industries. For microbial production of β-carotene in Saccharomyces cerevisiae, the supply of geranylgeranyl diphosphate (GGPP) was firstly increased in S. cerevisiae BY4742 to obtain strain BY4742-T2 through over-expressing truncated 3-hydroxy-3-methylglutaryl-CoA reductase (tHMGR), which is the major rate-limiting enzyme in the mevalonate (MVA) pathway, and GGPP synthase (GGPS), which is a key enzyme in the diterpenoid synthetic pathway. The β-carotene synthetic genes of Pantoea agglomerans and Xanthophyllomyces dendrorhous were further integrated into strain BY4742-T2 for comparing β-carotene production. Over-expression of tHMGR and GGPS genes led to 26.0-fold increase of β-carotene production. In addition, genes from X. dendrorhous was more efficient than those from P. agglomerans for β-carotene production in S. cerevisiae. Strain BW02 was obtained which produced 1.56 mg/g (dry cell weight) β-carotene, which could be used further for constructing cell factories for β-carotene production.