玉米黄烷酮-3-羟化酶原核表达条件对香橙素生成量的影响

Effect of prokaryotic expression conditions of Zea mays flavanone-3-hydroxylase on production of aromadendrin

香橙素是类黄酮代谢途径中重要的中间化合物,黄烷酮-3-羟化酶(flavanone-3-hydroxylase,F3H)能以柚皮素为底物催化C;位接羟基从而生成香橙素。为实现香橙素绿色发展,该研究利用微生物法,对来自玉米的Zmf3h构建大肠杆菌原核表达系统后诱导ZmF3H蛋白表达。通过发酵实验验证ZmF3H在胞内可溶性表达并成功催化生成香橙素。为提高香橙素产量,对蛋白表达阶段的菌体密度、异丙基-β-D-硫代半乳糖苷(isopropyl-β-D-thiogalactoside,IPTG)诱导剂浓度、诱导时间和诱导温度等条件进行优化实验。单因素试验结果表明各实验组分别在菌种密度OD;=0.8~1.0,IPTG终浓度为0.6 mmol/L,诱导温度在23℃,诱导8 h时可明显提高香橙素生成量,综合各单因素优化条件进行实验验证后发现香橙素生成量为(32.27±1.98)mg/L,与初始条件相比提高了60.39%。

Aromadendrin is an important intermediate compound in flavonoid metabolism pathway.Flavanone-3-hydroxylase(F3 H)catalyzes hydroxyl grafting of naringin at C;site.Microbial method was used to achieve the green development of aromadendrin.ZmF3 H protein was expressed by Escherichia coli containing Zmf3h from Zea mays.Fermentation experiments confirmed that ZmF3 H was soluble in intracellular and successfully catalyzed the production of aromadendrin.The results showed that the production of aromadendrin was significantly increased when the final IPTG concentration was 0.6 mmol/L,the induction temperature was 23℃,the induction time was 8 h,and the cell density was between 0.8 to 1.0.The production of aromadendrin was(32.27±1.98)mg/L after optimization which was 60.39%higher than that of initial conditions.