D-塔格糖3-差向异构酶的理性设计与机制解析  

Rational design and mechanism analysis of D-tagatose 3-epimerase

作　　者：郭丁钰 魏婉清 宋伟 吴静 闻建 胡贵鹏 刘立明 GUO Dingyu;WEI Wanqing;SONG Wei;WU Jing;WEN Jian;HU Guipeng;LIU Liming(School of Food Engineering,Anhui Science and Technology University,Chuzhou 233100,China;School of Biotechnology and Key Laboratory of Industrial Biotechnology of Ministry of Education,Jiangnan University,Wuxi 214122,China;School of Life Sciences and Health Engineering,Jiangnan University,Wuxi 214122,China)

机构地区：[1]安徽科技学院食品工程学院,安徽滁州233100 [2]江南大学生物工程学院,工业生物技术教育部重点实验室,江苏无锡214122 [3]江南大学生命科学与健康工程学院,江苏无锡214122

出　　处：《食品与发酵工业》2025年第4期58-64,共7页Food and Fermentation Industries

基　　金：国家重点研发计划项目(2021YFC2100100);中央高校基本科研业务费专项资金资助(JUSRP123011)。

摘　　要：D-阿洛酮糖是一种在自然界中存在极少的酮糖,因其具有与蔗糖相近的甜度但热量更低而备受关注。然而,D-阿洛酮糖的高效生物合成仍然面临挑战,是由于用于生产的酮糖3-差向异构酶的热稳定性较差。该研究通过理性设计,引入二硫键和属性嫁接策略,成功获得了热稳定性更高的最佳突变体M03。最佳突变体M03的T m值从52.7℃提高到69.4℃;在无需外源添加金属离子的条件下,55℃时半衰期延长至254.3 min,是野生型的54.1倍;通过分子动力学模拟和结构分析揭示了性能提升的详细机制。该研究为工业化生产D-阿洛酮糖提供了一个有潜力的突变菌株。D-allulose is a rare ketose found in nature,which has attracted significant attention for its similar sweetness to sucrose but lower caloric content.However,the efficient biosynthesis of D-allulose still poses a challenge,primarily attributed to the low thermostability of the ketose 3-epimerase utilized in production.In this study,the optimal mutant M03 with higher thermostability was developed by rational design,through the introduction of a disulfide bond and attribute grafting strategy.The T m value of the best mutant M03 increased from 52.7℃to 69.4℃.At 55℃,the half-life was extended to 254.3 min without metal ions,which was 54.1 times that of the wild-type.The detailed mechanism of performance improvement was analyzed by molecular dynamics simulation and structure analysis.This study provided a potential mutant strain for the industrial production of D-allulose.

关 键 词：D-阿洛酮糖 D-塔格糖3-差向异构酶 热稳定性 蛋白质工程改造 分子动力学模拟 

分 类 号：TS201.25[轻工技术与工程—食品科学]