Rosavin is the characteristic component of Rhodiola rosea L., an important medicinal plant used widely in the world that has been reported to possess multiple biological activities. However, the endangered status of wild Rhodiola has limited the supply of rosavin. In this work, we successfully engineered an Escherichia coli strain to efficiently produce rosavin as an alternative production method. Firstly, cinnamate: CoA ligase from Hypericum calycinum, cinnamoyl-CoA reductase from Lolium perenne, and uridine diphosphate (UDP)-glycosyltransferase (UGT) from Bacillus subtilis (Bs-YjiC) were selected to improve the titer of rosin in E. coli. Subsequently, four UGTs from the UGT91R subfamily were identified to catalyze the formation of rosavin from rosin, with SlUGT91R1 from Solanum lycopersicum showing the highest activity level. Secondly, production of rosavin was achieved for the first time in E. coli by incorporating the SlUGT91R1 and UDP-arabinose pathway, including UDP-glucose dehydrogenase, UDP-xylose synthase, and UDP-xylose 4-epimerase, into the rosin-producing stain, and the titer reached 430.5 ± 91.4 mg/L. Thirdly, a two-step pathway derived from L-arabinose, composed of L-arabinokinase and UDP-sugar pyrophosphorylase, was developed in E. coli to further optimize the supply of the precursor UDP-arabinose. Furthermore, 1203.7 ± 32.1 mg/L of rosavin was produced from D-glucose and L-arabinose using shake-flask fermentation. Finally, the production of rosavin reached 7539.1 ± 228.7 mg/L by fed-batch fermentation in a 5-L bioreactor. Thus, the microbe-based production of rosavin shows great potential for commercialization. This work provides an effective strategy for the biosynthesis of other valuable natural products with arabinose-containing units from D-glucose and L-arabinose.

中文翻译：

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在工程化大肠杆菌中从 D-葡萄糖和 L-阿拉伯糖中高水平生产红景天特征成分 rosavin


肉桂醇甙是红景天（Rhodiola rosea L.）的特征成分，红景天是一种在世界范围内广泛使用的重要药用植物，据报道具有多种生物活性。然而，野生红景天濒临灭绝的地位限制了肉桂醇甙的供应。在这项工作中，我们成功地设计了一种大肠杆菌菌株，以有效地生产 rosavin 作为替代生产方法。首先，选择肉桂酸酯：来自金丝翘的 CoA 连接酶、来自冻疮的肉桂酰辅酶 A 还原酶和来自枯草芽孢杆菌的尿苷二磷酸 （UDP）-糖基转移酶 （UGT） 以提高大肠杆菌中松香的滴度。随后，鉴定出来自 UGT91R 亚家族的 4 个 UGTs 催化松香形成肉桂醇甙，其中来自番茄的 SlUGT91R1 显示出最高的活性水平。其次，通过将 SlUGT91R1 和 UDP-阿拉伯糖途径（包括 UDP-葡萄糖脱氢酶、UDP-木糖合酶和 UDP-木糖 4-差向异构酶）掺入松香产染色剂中，首次在大肠杆菌中实现了肉桂醇素的产生，滴度达到 430.5 ± 91.4 mg/L。然后，在大肠杆菌中开发了源自 L-阿拉伯糖的两步途径，由 L-阿拉伯糖和 UDP-糖焦磷酸化酶组成，以进一步优化前体 UDP-阿拉伯糖的供应。 此外，使用摇瓶发酵从 D-葡萄糖和 L-阿拉伯糖中生产了 1203.7 ± 32.1 mg/L 的肉桂醇素。最后，在 5 L 生物反应器中通过补料分批发酵，rosavin 的产量达到 7539.1 ± 228.7 mg/L。因此，基于微生物的 rosavin 生产显示出巨大的商业化潜力。 这项工作为来自 D-葡萄糖和 L-阿拉伯糖的含阿拉伯糖单元的其他有价值的天然产物的生物合成提供了一种有效的策略。