Fang Lixia, Wen Peishi, Zhang Jiaqi, Cao Yingxiu
State Key Laboratory of Synthetic Biology, Tianjin University, Tianjin, China.
Frontiers Science Center for Synthetic Biology and Key Laboratory of Systems Bioengineering (Ministry of Education), School of Chemical Engineering and Technology, Tianjin University, Tianjin, China.
Biotechnol Bioeng. 2025 Sep;122(9):2499-2510. doi: 10.1002/bit.29041. Epub 2025 May 29.
Microbial production of free fatty acids (FFAs) is an eco-friendly and promising approach. However, FFAs accumulation within microbial cells imposes stress and toxicity, impairing biosynthetic performance. These challenges can be alleviated by exporting FFAs through the efflux system AcrAB-TolC in Escherichia coli, but the expression of these membrane channels needs to be fine-tuned. In this study, we employed a quorum sensing (QS) system to dynamically regulate the expression of AcrAB-TolC, thereby enhancing FFAs efflux and production. Two regulatory patterns of gene expression were designed to respond to increasing cell density: EsaI/R-P drives an initial increase followed by a decrease pattern, and EsaI/R-P enables a slow linear increase pattern. The L19IR-P-acrA strain, in which acrA expression is auto-induced by P, exhibited a 142% increase in extracellular FFAs and an 11% increase in total FFAs titer compared to the L19IR strain, which lacks acrA regulation. Auto-induced dynamic regulation of acrA demonstrates significant improvements in both FFAs efflux and overall production, while maintaining cell growth and membrane stability. Our results highlight the potential of QS-mediated dynamic regulation of efflux pumps to enhance the synthesis of bioproducts toxic to microorganisms.
微生物生产游离脂肪酸（FFAs）是一种环保且有前景的方法。然而，FFAs在微生物细胞内的积累会施加压力和毒性，损害生物合成性能。通过大肠杆菌中的外排系统AcrAB-TolC输出FFAs可以缓解这些挑战，但这些膜通道的表达需要进行微调。在本研究中，我们采用群体感应（QS）系统动态调节AcrAB-TolC的表达，从而增强FFAs的外排和生产。设计了两种基因表达调控模式以响应细胞密度的增加：EsaI/R-P驱动先增加后降低的模式，而EsaI/R-P实现缓慢线性增加的模式。与缺乏acrA调控的L19IR菌株相比，acrA表达由P自动诱导的L19IR-P-acrA菌株细胞外FFAs增加了142%，总FFAs滴度增加了11%。acrA的自动诱导动态调节在FFAs外排和总体生产方面均显示出显著改善，同时保持细胞生长和膜稳定性。我们的结果突出了QS介导的外排泵动态调节在增强对微生物有毒的生物产品合成方面的潜力。
