Efficient and directional supply of reducing equivalents is crucial for high-yield production of desired chemicals. Herein, an engineered isobutanol-producing Shewaenlla oneidensis was modularly constructed by assembling the electro-controlled distribution system of reducing equivalents and the isobutanol biosynthesis pathway. A dual-stage isobutanol electro-fermentation process was first established, including +0.5 V for cell growth and −0.6 V for isobutanol synthesis. Then, a redox biosensor-based dynamic regulation system was constructed to further decouple cell growth and isobutanol synthesis phases, enabling efficient supply of reducing equivalents. Lastly, an electro-controlled CRISPRi transcription inhibition system was designed to inhibit competitive metabolic pathways, which led to directional distribution of reducing equivalents and carbon flux toward isobutanol biosynthesis. Thus, the titer of isobutanol reached 1,321.5 ± 106.8 mg/L, a 10.8-fold increase from the original strain with 94.9% of the theoretical yield. This study achieved electro-controlled directional distribution of reducing equivalents and enhanced biosynthesis of reductive products via microbial electro-fermentation.

中文翻译：

---

还原当量的电控制分布以促进 S. oneidensis 微生物电发酵中异丁醇的生物合成


高效、定向地供应还原当量对于高产量生产所需化学品至关重要。在此，通过组装还原当量的电控分配系统和异丁醇生物合成途径，模块化构建了工程化的异丁醇生产 Shewaenlla oneidensis。首先建立了双级异丁醇电发酵工艺，包括用于细胞生长的 +0.5 V 和用于异丁醇合成的 −0.6 V。然后，构建了基于氧化还原生物传感器的动态调节系统，以进一步解耦细胞生长和异丁醇合成阶段，从而能够高效供应还原当量。最后，设计了一种电控 CRISPRi 转录抑制系统来抑制竞争性代谢途径，这导致还原当量和碳通量向异丁醇生物合成的方向分布。因此，异丁醇的效价达到 1,321.5 ± 106.8 mg/L，比原始菌株提高了 10.8 倍，理论产量为 94.9%。本研究通过微生物电发酵实现了还原当量的电控制定向分布和增强还原产物的生物合成。