唾液酸生物制造急需的非辅酶依赖型N-乙酰葡萄糖胺异构酶的分子催化机制研究

项目名称： 唾液酸生物制造急需的非辅酶依赖型N-乙酰葡萄糖胺异构酶的分子催化机制研究

项目编号： No.21306083

项目类型： 青年科学基金项目

立项/批准年度： 2014

项目学科： 化学工业

项目作者： 谢婧婧

作者单位： 南京工业大学

项目金额： 25万元

中文摘要： 唾液酸N-乙酰神经氨酸（Neu5Ac）及其类似物具有广泛的生理功能，在医药、食品等行业中应用广泛，国内外的市场需求极大。目前采用N-乙酰葡萄糖胺异构酶Age与N-乙酰神经氨酸醛缩酶Nal大规模合成Neu5Ac。但已报道的Age酶均需辅酶ATP激活，这是制约Neu5Ac低成本、高效生物制造的瓶颈。本课题组自主发掘了目前唯一一个非辅酶依赖型的N-乙酰葡萄糖胺异构酶PmAge。本项目拟通过建立PmAge酶分子的三维结构模型，分析其功能结构域的特征和性质，获得PmAge以底物N-乙酰葡萄糖胺在非辅酶ATP依赖下异构生成N-乙酰甘露糖胺的催化机制。运用该机理指导构建非辅酶依赖的Ch1Age突变酶，为理性设计消除此类辅酶激活提供相关共性方法。本项目中研究的PmAge与Ch1Age突变酶，也将成为在工业生物制造Neu5Ac中具有应用潜力的生物催化剂。

中文关键词： N-乙酰葡萄糖胺异构酶；N-乙酰神经氨酸醛缩酶；N-乙酰神经氨酸；构效关系；催化机制

英文摘要： Sialic acid Neu5Ac and its analogues have a wide range of physiological functions. Its applications in medicine, food and other industries prospect a great market demand domestic and abroad. N-acetylglucosamine 2-epimerase (Age) and N-acetylneuraminic acid lyase (Nal) are catalysts in biological manufacturing of Neu5Ac. The request of coenzyme ATP of all reported Ages is the bottleneck of low-cost and efficient bio-manufacturing of Neu5Ac. Our group discovered the only cofactor independent N-acetylglucosamine 2-epimerase, named PmAge. In this research object, we would like to create a homology model of the PmAge molecule, analyze the characteristics and properties of its functional domains, research the heterogeneous catalytic mechanism how PmAGE isomerize N-acetyl glucosamine to N-acetylmannosamine without co-activation of ATP. Guided by the catalytic mechanism, we would like to construct cofactor independent Ch1Age mutants. This process would provide relevant common methods for the rational design of elimination of cofactor. PmAge and Ch1Age mutants generated in this project can also be used as the candidate catalysts to promote existing Neu5Ac industrial biomanufacturing.

英文关键词： N-acetylglucosamine isomerase；N-acetylneuraminic acid aldolase；N-acetylneuraminic acid；Structure-function relationship；Catalytic mechanism