代谢工程改造毕赤酵母生产葡萄糖二酸  

Metabolic Engineering of Pichia pastoris for Production of Glucaric Acid

作　　者：刘叶[1,2] 巩旭[1,2] 康振[1,2] 堵国成[1,2] 陈坚[1,2] 华兆哲[1,2] LIU Ye;GONG Xu;KANG Zheng;DU Guocheng;CHEN Jian;HUA Zhaozhe(Key Laboratory of Industrial Biotechnology,Ministry of Education,Jiangnan University,Wuxi 214122,China;School of Biotechnology,Jiangnan University,Wuxi 214122,China)

机构地区：[1]江南大学工业生物技术教育部重点实验室,江苏无锡214122 [2]江南大学生物工程学院,江苏无锡214122

出　　处：《食品与生物技术学报》2018年第9期955-961,共7页Journal of Food Science and Biotechnology

基　　金：国家973计划项目(2013CB733602);国家863计划项目(2011AA100905);江苏省博士后基金项目(1301010B);教育部长江学者和创新团队发展计划项目(IRT1135)

摘　　要：葡萄糖二酸是一种具有重要生物功能的有机酸,为了实现微生物合成葡萄糖二酸的高效生产,异源表达来源于小鼠基因组的MIOX基因及Pseudomonas putida KT2440的udh基因,在毕赤酵母中构建葡萄糖二酸合成途径,实现了葡萄糖二酸的生成。通过在摇瓶水平进行优化,确定了最适碳源为葡萄糖(初始质量浓度60 g/L最优),最适初始pH为5.5,最优接种龄为24 h,最适接种体积分数为25%,重组毕赤酵母生成的葡萄糖二酸产量从最初的(566.36±16.98) mg/L提高至(967.60±3.90) mg/L。在此基础上,重组毕赤酵母在3 L发酵罐中放大培养,葡萄糖二酸的产量达到了(2.60±0.04) g/L。Glucaric acid is a kind of organic acid with important biological values.In order to achieve high-yield production of glucaric acid,a glucaric acid synthetic pathway was constructed in Pichia pastoris by co-expression of the mouse MIOX gene and the udh gene from Pseudomonas putida KT2440,which led to accumulation of glucaric acid.By fermentation optimization in shake flasks,the results showed the optimum carbon source was glucose(60 g/L was the best initial concentration),the optimum initial pH was 5.5,the optimum seed time was 24 h,and the optimum inoculation amount was 25%.The glucaric acid production was increased from(566.36±16.98)mg/L to(967.60±3.90)mg/L after optimization.According to the optimized conditions,the production of glucaric acid was substantially increased to(2.60±0.04)g/L in a 3 L fermentor.

关 键 词：葡萄糖二酸 毕赤酵母 代谢工程 发酵优化 

分 类 号：Q815[生物学—生物工程]