Zhu Zhangliang, Li Chao, Liu Xin, Gao Dengke, Wang Xueyu, Tanokura Masaru, Qin Hui-Min, Lu Fuping
Key Laboratory of Industrial Fermentation Microbiology of the Ministry of Education, Tianjin Key Laboratory of Industrial Microbiology, College of Biotechnology, Tianjin University of Science and Technology, National Engineering Laboratory for Industrial Enzymes Tianjin 300457 P. R. China
Laboratory of Basic Science on Healthy Longevity, Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo 1-1-1 Yayoi, Bunkyo Tokyo 113-8657 Japan
RSC Adv. 2019 Jan 22;9(6):2919-2927. doi: 10.1039/c8ra10029b.
sp. d-tagatose 3-epimerase (sDTE) catalyzes the conversion of d-tagatose to d-sorbose. It also recognizes d-fructose as a substrate for d-allulose production. The optimal temperature and pH of the purified sDTE was 50 °C and 8.0, respectively. Based on the sDTE homologous model, Glu154, Asp187, Gln213, and Glu248, form a hydrogen bond network with the active-site Mn and constitute the catalytic tetrad. The amino acid residues around O-1, -2, and -3 atoms of the substrates (d-tagatose/d-fructose) are strictly conserved and thus likely regulate the catalytic reaction. However, the residues at O-4, -5, and -6, being responsible for the substrate-binding, are different. In particular, Arg65 and Met9 were found to form a unique interaction with O-4 of d-fructose and d-tagatose. The whole cells with recombinant sDTE showed a higher bioconversion rate of 42.5% in a fed-batch bioconversion using d-fructose as a substrate, corresponding to a production of 476 g Ld-allulose. These results suggest that sDTE is a potential industrial biocatalyst for the production of d-allulose in fed-batch mode.
D-塔格糖3-表异构酶（sDTE）催化D-塔格糖转化为D-山梨糖。它还识别D-果糖作为生产D-阿洛酮糖的底物。纯化后的sDTE的最适温度和pH分别为50℃和8.0。基于sDTE同源模型，Glu154、Asp187、Gln213和Glu248与活性位点的锰形成氢键网络并构成催化四联体。底物（D-塔格糖/D-果糖）的O-1、-2和-3原子周围的氨基酸残基严格保守，因此可能调节催化反应。然而，负责底物结合的O-4、-5和-6处的残基不同。特别是，发现Arg65和Met9与D-果糖和D-塔格糖的O-4形成独特的相互作用。在以D-果糖为底物的补料分批生物转化中，含有重组sDTE的全细胞显示出42.5%的更高生物转化率，对应于476 g/L D-阿洛酮糖的产量。这些结果表明，sDTE是补料分批模式下生产D-阿洛酮糖的潜在工业生物催化剂。
