Abstract：
Vanillin is one of the world\'s most extensively used flavoring agents with high application value. However, the yield of vanillin biosynthesis remains limited due to the low efficiency of substrate uptake and the inhibitory effect on cell growth caused by vanillin. Here, we screened high-efficiency ferulic acid importer TodX and vanillin exporters PP_0178 and PP_0179 by overexpressing genes encoding candidate transporters in a vanillin-producing engineered Escherichia coli strain VA and further constructed an autoregulatory bidirectional transport system by coexpressing TodX and PP_0178/PP_0179 with a vanillin self-inducible promoter ADH7. Compared with strain VA, strain VA-TodX-PP_0179 can efficiently transport ferulic acid across the cell membrane and convert it to vanillin, which significantly increases the substrate utilization rate efficiency (14.86%) and vanillin titer (51.07%). This study demonstrated that the autoregulatory bidirectional transport system significantly enhances the substrate uptake efficiency while alleviating the vanillin toxicity issue, providing a promising viable route for vanillin biosynthesis.
摘要：
香兰素是世界上应用最广泛的调味剂之一,具有很高的应用价值。然而,香兰素生物合成的产量仍然有限，由于底物吸收效率低，以及香兰素对细胞生长的抑制作用。这里,我们通过在生产香兰素的工程大肠杆菌VA菌株中过表达编码候选转运蛋白的基因，筛选了高效阿魏酸进口蛋白TodX和香兰素出口商PP_0178和PP_0179，并通过共表达TodX和PP_0178/PP_0179进一步构建了自动调节双向运输系统。香兰素自诱导型启动子ADH7。与VA菌株相比，菌株VA-TodX-PP_0179可以有效地将阿魏酸穿过细胞膜并将其转化为香草醛，显著提高了底物利用率(14.86%)和香草醛效价(51.07%)。这项研究表明，自动调节双向运输系统显着提高底物吸收效率，同时减轻香草醛毒性问题，为香草醛生物合成提供了一条有前途的可行路线。
