高山被孢霉ω-6/ω-3脂肪酸代谢流调控"分子开关"的分子改造及其机制研究

项目名称： 高山被孢霉ω-6/ω-3脂肪酸代谢流调控"分子开关"的分子改造及其机制研究

项目编号： No.31271812

项目类型： 面上项目

立项/批准年度： 2013

项目学科： 农业科学

项目作者： 宋元达

作者单位： 江南大学

项目金额： 77万元

中文摘要： 高山被孢霉是商业化生产ω-6脂肪酸ARA的重要菌种，但也能合成极少量的ω-3脂肪酸EPA。ARA/EPA合成通路由Δ12 Des、ω3 Des、Δ6 Des、Δ6 Elo和Δ5 Des组成。其中处于代谢流分支点的Δ6 Des和ω3 Des是调控ω-6/ω-3脂肪酸代谢流的"分子开关"。但我们对该分子开关如何通过其底物偏好性和活性来调节脂肪酸代谢流的分子机制还知之甚少。本课题将采用点饱和突变技术对分子开关进行分子改造，分析突变体的活性、底物偏好性与分子结构的相互关系；并通过体内实验对其进行功能验证；探索ω-6/ω-3脂肪酸代谢流的调控机制；最后构建高产EPA细胞工厂。本课题将为高山被孢霉工业化生产EPA或ω-6/ω-3比例为1:1的营养平衡的膳食油脂奠定技术基础，也将扩展我们对微生物选择性合成ω-6/ω-3 脂肪酸的生物化学基础的认识。

中文关键词： 高山被孢霉；分子开关；Δ6脱氢酶；ω-3脱氢酶；二十碳五烯酸

英文摘要： As a key commercial producer of ω-6 fatty acid, arachidonic acid (ARA), Mortierella alpina can also produce a little amount of ω-3 fatty acid,eicosapentaenoic acid (EPA).The key enzymes involved in ARA/EPA biosynthetic pathway are: Δ12 Des、ω3 Des、Δ6 Des、Δ6 Elo andΔ5 Des.Among which Δ6 Des and ω3 Des,located at the branch point of fatty acid metabolic flux,are key regulators of ω-6/ω-3 fatty acid metabolic flux.These two enzymes work together as a molecular switch that controlω-6/ω-3 metabolic flux. The molecular switch may regulate the metabolic flux through its activity and substrate preference, but up to present,molecular base of the function of the molecular switch is unclear. In this project, we will investigate the relationship between the structure and activity and substrate preference using saturated site-directed mutagenesis technique, and this will be further confirmed by in vivo study.We will also try to crystallize the Δ6 Des, and solve its crystal structure.If we can solve the structure of Δ6 Des, or other scientists may solve any membrane bound desaturase, then we can further explore the structure and function of the molecular switch。This work will provide technical support to develop an EPA or ω-6/ω-3 ratio balanced oil cell factorty, and will also provide insights into the biochemistry of differen

英文关键词： Mortierella alpina；Molecular switch；Δ6 dehydrogenase；ω-3 dehydrogenase；EPA