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食品与发酵工业  2021, Vol. 47 Issue (21): 79-85    DOI: 10.13995/j.cnki.11-1802/ts.026301
  研究报告 本期目录 | 过刊浏览 | 高级检索 |
甜橙来源法尼烯合成酶的异源表达及酶学性质研究
付闻文1,2, 王均华1,2, 李由然1,2, 石贵阳1,2*
1(粮食发酵工艺与技术国家工程实验室(江南大学),江苏 无锡,214122)
2(江南大学 生物工程学院,江苏 无锡,214122)
Heterologous expression and enzymatic properties of farnesene synthase from Citrus sinensis
FU Wenwen1,2, WANG Junhua1,2, LI Youran1,2, SHI Guiyang1,2*
1(National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi 214122, China)
2(School of Biotechnology, Jiangnan University, Wuxi 214122, China)
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摘要 法尼烯是一种倍半萜化合物,在食品、香料等领域有广泛应用价值。法尼烯合成酶是法尼烯合成途径中的关键限速酶,对法尼烯的合成有重要催化作用。通过生物信息学方法获取了甜橙来源萜类合成酶(farnesene synthase from Citrus sinensis,FSCS)基因Fscs, 通过异源表达对其进行了功能鉴定,研究了重组酶的酶学性质并将其应用于酿酒酵母法尼烯合成。结果显示,在以法尼基焦磷酸为底物时,FSCS可催化其生成α-法尼烯;Mg2+为酶必需的辅因子,K+可促进产物生成,8 mmol/L以上Mn2+对产物生成有抑制作用,最适金属离子条件为30 mmol/L Mg2+, 40 mmol/L K+;最适反应温度为20 ℃,酶对高温耐受性差。将酶学性质应用于酿酒酵母重组表达菌株,最适金属离子条件下法尼烯产量提升了30%,达到43.76 mg/L;温度条件的改变对发酵后期法尼烯积累有一定促进作用。该研究对一种新的萜类合成酶进行了功能表征,并通过酶学性质研究为构建微生物细胞工厂高效生产法尼烯提供了参考。
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付闻文
王均华
李由然
石贵阳
关键词:  法尼烯合成酶  酶学性质  法尼基焦磷酸  倍半萜  酿酒酵母    
Abstract: Farnesene is a sesquiterpene compound which has a wide range of applications in food, fragrance & flavor and other industries. Farnesene synthase is the key enzyme in biosynthesis of farnesene. Its properties significantly affect the formation of farnesene. A farnesene synthase gene from Citrus sinensis (FSCS) was cloned and expressed in Escherichia coli BL21(DE3). The recombinant enzyme was purified and its biochemical characteristics were investigated. Results showed that FSCS could produce α-farnesene by using farnesyl diphosphate as substrate. In addition, Mg2+ was indispensable, and K+ could enhance the activity of FSCS. However, 8 mmol/L or more Mn2+ significantly inhibited its activity. The optimal condition of Mg2+ and K+ were 30 mmol/L and 40 mmol/L, respectively. The optimal temperature was 20℃, and sensitive to higher temperature. Under the optimal conditions in recombinant Saccharomyces cerevisiae, the yield of farnesene was 43.76 mg/L increased by 30%. The results could provide reference for efficient production of farnesene in microbial cell factories.
Key words:  farnesene synthase    characteristics    farnesyl diphosphate    sesquiterpene    Saccharomyces cerevisiae
收稿日期:  2020-11-27      修回日期:  2021-03-10           出版日期:  2021-11-15      发布日期:  2021-11-30      期的出版日期:  2021-11-15
基金资助: 国家重点研发计划项目(2018YFA0900504)
作者简介:  付闻文硕士研究生和王均华博士研究生为共同第一作者(石贵阳教授为通讯作者,E-mail:gyshi@jiangnan.edu.cn)
引用本文:    
付闻文,王均华,李由然,等. 甜橙来源法尼烯合成酶的异源表达及酶学性质研究[J]. 食品与发酵工业, 2021, 47(21): 79-85.
FU Wenwen,WANG Junhua,LI Youran,et al. Heterologous expression and enzymatic properties of farnesene synthase from Citrus sinensis[J]. Food and Fermentation Industries, 2021, 47(21): 79-85.
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http://sf1970.cnif.cn/CN/10.13995/j.cnki.11-1802/ts.026301  或          http://sf1970.cnif.cn/CN/Y2021/V47/I21/79
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