Yeast productivity in lignocellulosic ethanol fermentation is clearly impeded by stress. Enhancing the robustness of xylose-fermenting yeast is important for improving lignocellulosic ethanol production. In this study, the glutathione biosynthesis pathway and acetic acid degradation pathway were strengthened to enhance yeast tolerance to stress due to elevated reactive oxygen species (ROS) and acetic acid. Dynamic feedback regulation of the anti-stress genetic circuits was achieved using stress-driven promoters discovered from the transcriptome to maintain low intracellular ROS, relieve the metabolic burden, and ultimately improve the robustness and ethanol production of yeast. The cell growth, xylose utilization and ethanol production of the engineered strain were enhanced under both stress and nonstress conditions. The engineered strain showed 49.5% and 17.5% higher ethanol productivity in laboratory media and industrial lignocellulosic media, respectively, at 36 °C compared with the parent strain. This study provides novel insights on the rational design and construction of feedback genetic circuits for dynamically improving yeast robustness.

木质纤维素乙醇发酵中的酵母生产力明显受到压力的阻碍。提高木糖发酵酵母的稳健性对于提高木质纤维素乙醇的生产很重要。在这项研究中，谷胱甘肽生物合成途径和乙酸降解途径得到加强，以增强酵母对活性氧 (ROS) 和乙酸升高引起的应激的耐受性。使用从转录组中发现的应激驱动启动子实现抗应激遗传回路的动态反馈调节，以维持低细胞内 ROS，减轻代谢负担，并最终提高酵母的健壮性和乙醇产量。工程菌株的细胞生长、木糖利用和乙醇产量在胁迫和非胁迫条件下均得到增强。与亲本菌株相比，在 36 °C 下，工程菌株在实验室培养基和工业木质纤维素培养基中的乙醇生产率分别提高了 49.5% 和 17.5%。这项研究为反馈遗传电路的合理设计和构建提供了新的见解，以动态提高酵母的稳健性。