Metabolic Engineering of a curdlan-producing Agrobacterium sp. for sugar nucleotide cofactor regeneration

产凝胶多糖农杆菌的代谢工程。

• 批准号: 0233753
• 负责人: Ruizhen Chen
• 金额: $ 23.29万
• 依托单位: Virginia Commonwealth University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-01 至 2004-10-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0233753&HistoricalAwards=false
• 关键词: Metabolic; Engineering; curdlan; producing; Agrobacterium

Agrobacterium sp. ATCC 31749 is an industrial strain used to produce curdlan, a homo-glucose polymer. The unusually high productivity of the glucose polymer in this strain results from a highly efficient UDP-Glucose cofactor regeneration mechanism. This project proposes to use this naturally evolved mechanism as a platform to develop highly efficient sugar nucleotide cofactor regeneration systems useful for enzymatic oligosaccharide synthesis. First, the polymerization step can be disrupted by introducing mutations in the curdlan synthase gene. The resulting mutant can function as an efficient UDP- glucose regeneration system. Further, taking advantage of the fact that UDP-glucose is the entry point of the biosynthetic pathways of three other sugar nucleotides, the research plan proposes strategies to transform the genetically modified Agrobacterium strain into UDP-galactose, UDP-glucuronic acid and UDP-galacuronic acid regeneration systems. This project will also contribute to metabolic engineering/biocatalysis research through new and effective genetic and broad-host-range based heterologous protein expression methods applicable to a wide range of non-E.coli bacteria.

农杆菌ATCC 31749是用于生产可得兰（一种高葡萄糖聚合物）的工业菌株。该菌株中葡萄糖聚合物的异常高的生产率是由高效的UDP-葡萄糖辅因子再生机制引起的。该项目提出使用这种自然进化的机制作为平台，开发用于酶促寡糖合成的高效糖核苷酸辅因子再生系统。首先，聚合步骤可以通过在可德兰合酶基因中引入突变来破坏。所得到的突变体可以作为一个有效的UDP-葡萄糖再生系统。此外，利用UDP-葡萄糖是其他三种糖核苷酸生物合成途径的切入点这一事实，研究计划提出了将遗传修饰的农杆菌菌株转化为UDP-半乳糖、UDP-葡萄糖醛酸和UDP-半乳糖醛酸再生系统的策略。该项目还将通过适用于各种非大肠杆菌细菌的新的有效的遗传和广泛宿主范围的异源蛋白表达方法，为代谢工程/生物催化研究做出贡献。