过表达 EHT1 基因对酿酒酵母己酸乙酯生产能力的影响  

Effects of Overexpressed EHT1 on Ester Production of Saccharomyces cerevisiae

作　　者：李锋[1] 陈叶福[1] 郭建[1] 郭学武[1] 肖冬光[1] 

机构地区：[1]天津科技大学生物工程学院,工业发酵微生物教育部重点实验室,天津300457

出　　处：《现代食品科技》2014年第3期93-98,176,共7页Modern Food Science and Technology

基　　金：长江学者和创新研究团队项目(IRT1166);国家自然科学基金项目(31271916);“十二五”农村领域国家科技计划课题(2013AA102108);天津市科委青年基金资助(12JCQNJC06500)

摘　　要：为了提高酿酒酵母己酸乙酯的产量,缩短浓香型白酒的发酵周期,降低生产粮耗。对酿酒酵母醇己酰基转移酶(ethanol hexanoyl transferase I)基因EHT1进行了过表达,研究其对酿酒酵母己酸乙酯生产性能的影响。以大肠杆菌-酿酒酵母穿梭质粒YEp352为载体,磷酸甘油酸激酶基因PGK1启动子为上游调控元件,KanMX抗性基因为筛选标记构建了酵母菌表达质粒Yep-PEK,经醋酸锂转化和G418抗性筛选鉴定获得过表达EHT1基因的突变株AY-PEK。经玉米原料液态白酒发酵后,实验结果显示突变株的生长速度、发酵速度、酒精度等基本发酵性能没有明显变化,但己酸乙酯的产量提高为原菌种的2.21倍,辛酸乙酯和癸酸乙酯也分别提高了31.4%和49.1%。当加入等量的己酸模仿实际发酵过程中己酸菌提供己酸时,己酸乙酯的量提高为原菌种的近3倍,总酯也相应的提高了32.2%。EHT1基因的过表达对提高酿酒酵母产酯性能,特别是产己酸乙酯有显著作用。To increase the production of ethyl caproate by Chinese liquor yeast （S. cerevisiae AY15）, shorten the fermentation period, and reduce the grainconstunption, a recombinant strain AY-PEK was constructed by overexpressing EHT1 （encoding ethanol hexanoyl transferase） to investigate the effect of EHTl-overexpression on the performance of ester-producing. The EHT1 gene was amplified from genomic DNA of AY15, the PGK1 promoter and resistance gene KanMXwere orderly inserted into vector Yep352 to construct the expression plasmid Yep-PEK. Then EHTl-overexpression mutants AY-PEK were selected by YEPD agar plates containing 1400 ~tg/mL G418 after the plasmid Yep-PEK was transformed into AY15. The results showed that after liquid fermentation by corn, the differences in fermentation performances （growth rate, fermentation rate and the yield of alcohol） between AY-PEK and AY 15were negligible. However, the production of ethyl caproate produced by AY-PEK was markedly increased to 2.21-fold as high as that of parental strain AY15. Ethyl octanoate and ethyl decanoate were improved by 31.4% and 49.1%, respectively. The same amount of caproic acid was added to fermentation medium to imitate the actual fermentation, the production of ethyl caproate produced by AY-PEK was increased to nearly 2-fold higher than that of AY15, and the amount of total esters were also increased by 32.2%. Overexpression ofEHT1 gene can significantly enhance the production of ethyl esters, especially ethyl caproate.

关 键 词：中国白酒 酿酒酵母 己酸乙酯 醇己酰基转移酶 

分 类 号：TS261.11[轻工技术与工程—发酵工程]