Biosynthesis of (+)-catechin glycosides using recombinant amylosucrase from Deinococcus geothermalis DSM 11300
Biosynthesis of (+)-catechin glycosides using recombinant amylosucrase from Deinococcus geothermalis DSM 11300
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DOI：
10.1016/j.enzmictec.2011.05.007
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发表时间：
2011-07-10
影响因子：
3.4
通讯作者：
Park, Cheon-Seok
中科院分区：
文献类型：
--
作者：
Cho, Hyun-Kug;Kim, Hee-Hang;Park, Cheon-Seok
淀粉蔗糖酶（ASase，EC 2.4.1.4）是一种葡糖基转移酶，可将蔗糖水解为葡萄糖和果糖，并从释放的葡萄糖产生直链淀粉样葡聚糖聚合物。 (+)-儿茶素是一种植物多酚代谢物，具有美白和抗氧化活性。在本研究中，来自地热奇球菌 (dgas) 的 ASase 基因在大肠杆菌中表达，而重组 DGAS 酶则使用谷胱甘肽 S-转移酶融合系统进行纯化。利用 DGAS 转糖基活性从 (+)-儿茶素合成 (+)-儿茶素糖苷衍生物。我们使用 TLC 确认了两种主要转糖基产物的存在。 (+)-儿茶素转糖基化产物使用硅胶开放柱色谱和回收HPLC分离。通过 H-1 和 C-13 NMR 确定两种 (+)-儿茶素主要转移产物为 (+)-儿茶素-3'-O-α-D-吡喃葡萄糖苷，其中葡萄糖分子与 (+)-儿茶素连接，以及 (+)-儿茶素-3'-O-α-D-麦芽糖苷，其中麦芽糖与 (+)-儿茶素连接。通过回收 HPLC 和酶分析也证实了 DGAS 反应中存在 (+)-儿茶素麦芽低聚糖。研究了不同反应条件（温度、酶浓度、受体和供体摩尔比）对(+)-儿茶素糖苷的产率和类型的影响。 (C) 2011 Elsevier Inc. 保留所有权利。
Amylosucrase (ASase, EC 2.4.1.4) is a glucosyltransferase that hydrolyzes sucrose into glucose and fructose and produces amylose-like glucan polymers from the released glucose. (+)-Catechin is a plant polyphenolic metabolite having skin-whitening and antioxidant activities. In this study, the ASase gene from Deinococcus geothermalis (dgas) was expressed in Escherichia coli, while the recombinant DGAS enzyme was purified using a glutathione S-transferase fusion system. The (+)-catechin glycoside derivatives were synthesized from (+)-catechin using DGAS transglycosylation activity. We confirmed the presence of two major transglycosylation products using TLC. The (+)-catechin transglycosylation products were isolated using silica gel open column chromatography and recycling-HPLC. Two (+)-catechin major transfer products were determined through H-1 and C-13 NMR to be (+)-catechin-3'-O-alpha-D-glucopyranoside with a glucose molecule linked to (+)-catechin and (+)-catechin-3'-O-alpha-D-maltoside with a maltose linked to (+)-catechin. The presence of (+)-catechin maltooligosaccharides in the DGAS reaction was also confirmed via recycling-HPLC and enzymatic analysis. The effects of various reaction conditions (temperature, enzyme concentration, and molar ratio of acceptor and donor) on the yield and type of (+)-catechin glycosides were investigated. (C) 2011 Elsevier Inc. All rights reserved.