利用代谢工程在酿酒酵母中高效合成2-萘乙醇

Efficient Biosynthesis of 2-Naphthaleneethanol in Metabolically Engineered Saccharomyces cerevisiae

【目的】为实现生物合成重要化合物2-萘乙醇,探索利于2-萘乙醇合成的基因组合。【方法】向酿酒酵母BY4741发酵液中外源添加2-萘丙氨酸,运用高效液相色谱法进行产物检测;绘制BY4741的生长曲线和产量曲线,添加不同浓度2-萘丙氨酸到BY4741发酵液中并检测BY4741生长和生产情况;对BY4741进行代谢工程改造,构建过表达Ehrlich途径6个基因的质粒并转化进BY4741中;发酵6株改造菌株,检测产物产量。【结果】发酵液中检测到了推测产物2-萘乙醇,代谢工程改造菌株相较于野生型BY4741菌株产量均有所提升,其中过表达ARO9和ARO10基因使得2-萘乙醇的产量在外源添加1 mmol/L 2-萘丙氨酸的情况下达到0.258 mmol/L,转化率达到野生型的2.3倍。【结论】在酿酒酵母中实现了2-萘乙醇的生物合成,探索出ARO9和ARO10是利于2-萘乙醇生物合成的基因组合,为工业生产2-萘乙醇提供了一种新的生物合成方法。

【Objective】To elucidate the biosynthesis of 2-naphthaleneethanol and explore the gene combination beneficial to the synthesis of 2-naphthaleneethanol.【Method】2-Naphthylalanine was added to the fermentation broth of Saccharomyces cerevisiae BY4741 and the product was detected by HPLC.The growth curve and yield curve of BY4741 were explored.After adding different concentrations of 2-naphthylalanine into the fermentation broth of BY4741,the growth and production level were detected.The plasmids overexpressing six genes of Ehrlich pathway were constructed and transformed into BY4741.Six modified strains were fermented and the product yield was detected.【Result】2-Naphthaleneethanol was detected in the fermentation broth as expected.Compared with the wild type BY4741,the yields of the metabolically engineered strains were all improved.Overexpressing ARO9 and ARO10 made the yield of 2-naphthaleneethanol reached 0.258 mmol/L when exogenous 2-naphthylalanine was added,and the transformation rate reached 2.3 times that of the wild type.【Conclusion】The biosynthesis of 2-naphthaleneethanol was achieved in S.cerevisiae.ARO9 and ARO10 were found to be the optimal gene combinations beneficial to the biosynthesis of 2-naphthaleneethanol.This study provides a new biosynthesis method for industrial production of 2-naphthaleneethanol.