文献类型： 外文期刊
作者： Li, Lei 1 ; Zhang, Qianqian 1 ; Wang, Tong 1 ; Qi, Hongbin 1 ; Wei, Meijing 1 ; Lu, Fuping 1 ; Guan, Lijun 4 ; Mao, Shuhong 1 ; Qin, Hui-Min 1 ;
作者机构： 1.Tianjin Univ Sci & Technol, Key Lab Ind Fermentat Microbiol, Minist Educ, Tianjin 300457, Peoples R China
2.Tianjin Univ Sci & Technol, Coll Biotechnol, Tianjin Key Lab Ind Microbiol, Tianjin 300457, Peoples R China
3.Natl Engn Lab Ind Enzymes, Tianjin 300457, Peoples R China
4.Heilongjiang Acad Agr Sci, Inst Food Proc, Harbin 150086, Peoples R China
关键词： d-allulose; d-allulose 3-epimerase; protein engineering; acid resistance; functional juice
期刊名称：JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY （ 影响因子：5.895； 五年影响因子：6.048 ）
ISSN： 0021-8561
年卷期：
页码：
收录情况： SCI
摘要： D-Allulose, a rare sugar and functional sweetener, can be biosynthesized by D-allulose 3-isomerase (DAE). However, most of the reported DAEs exhibit poor resistance under acidic conditions, which severely limited their application. Here, surface charge engineering and random mutagenesis were used to construct a mutant library of CcDAE from Clostridium cellulolyticum H10, combined with high-throughput screening to identify mutants with high activity and resistance under acidic conditions. The mutant M3 (I114R/K123E/H209R) exhibited high activity (3.36-fold of wild-type) and acid resistance (10.6-fold of wild-type) at pH 4.5. The structure-function relationship was further analyzed by molecular dynamics (MD) simulations, which indicated that M3 had a higher number of hydrogen bonds and negative surface charges than the wild type. A multienzyme cascade system including M3 was used to convert high-calorie sugars in acidic juices, and functional juices containing 7.8-15.4 g/L D-allulose were obtained. Our study broadens the manufacture of functional foods containing D-allulose.
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