Improved Expression of His6‐Tagged Strictosidine Synthase cDNA for Chemo‐Enzymatic Alkaloid Diversification
Improved Expression of His6‐Tagged Strictosidine Synthase cDNA for Chemo‐Enzymatic Alkaloid Diversification
复制标题
DOI：
10.1002/cbdv.201000052
复制
发表时间：
2010-04
影响因子：
2.9
通讯作者：
Liuqing Yang;H. Zou;Huajian Zhu;M. Ruppert;Jingxu Gong;J. Stöckigt
中科院分区：
文献类型：
--
作者：
Liuqing Yang;H. Zou;Huajian Zhu;M. Ruppert;Jingxu Gong;J. Stöckigt
Structosidine synthase（STR 1）催化3α（S）-structosidine从色胺和裂叶甘露醇的立体选择性形成。阿托西啶是2，000种植物单萜吲哚生物碱生物合成的关键中间体，也是酶介导合成生物碱的关键前体。描述了一种改进的表达系统，其导致在大肠杆菌中优化的His 6-STR 1合成。通过确定伴侣蛋白pG-Tf 2和pG-LJE 8的共表达的影响来实现STR 1的最佳生产。在单独的伴侣蛋白pG-Tf 2存在下，STR 1的量和活性加倍。固定在Ni-NTA上的His 6-STR 1可用于制备规模的异胡桃树苷酶合成。与新的共表达的His 6-STR 1一起，通过酶促催化7-氮杂色胺和裂殖甘素获得新的3α（S）-12-氮杂木苷。所获得的结果对于应用化学-酶促方法导致具有新的改进结构的生物碱多样化具有重要意义。
Strictosidine synthase (STR1) catalyzes the stereoselective formation of 3α(S)‐strictosidine from tryptamine and secologanin. Strictosidine is the key intermediate in the biosynthesis of 2,000 plant monoterpenoid indole alkaloids, and it is a key precursor of enzyme‐mediated synthesis of alkaloids. An improved expression system is described which leads to optimized His6‐STR1 synthesis in Escherichia coli. Optimal production of STR1 was achieved by determining the impact of co‐expression of chaperones pG‐Tf2 and pG‐LJE8. The amount and activity of STR1 was doubled in the presence of chaperone pG‐Tf2 alone. His6‐STR1 immobilized on Ni‐NTA can be used for enzymatic synthesis of strictosidines on a preparative scale. With the newly co‐expressed His6‐STR1, novel 3α(S)‐12‐azastrictosidine was obtained by enzymatic catalysis of 7‐azatryptamine and secologanin. The results obtained are of significant importance for application to chemo‐enzymatic approaches leading to diversification of alkaloids with novel improved structures.