Bacteroides thetaiotaomicron represents a major symbiont of the human gut microbiome that is increasingly viewed as a promising candidate strain for microbial therapeutics. Here, we engineer B. thetaiotaomicron for heterologous production of non-native butyrate as a proof-of-concept biochemical at therapeutically relevant concentrations. Since B. thetaiotaomicron is not a natural producer of butyrate, we heterologously expressed a butyrate biosynthetic pathway in the strain, which led to the production of butyrate at the final concentration of 12 mg/L in a rich medium. Further optimization of butyrate production was achieved by a round of metabolic engineering guided by an expanded genome-scale metabolic model (GEM) of B. thetaiotaomicron. The in silico knock-out simulation of the expanded model showed that pta and ldhD were the potent knock-out targets to enhance butyrate production. The maximum titer and specific productivity of butyrate in the pta-ldhD double knockout mutant increased by nearly 3.4 and 4.8 folds, respectively. To our knowledge, this is the first engineering attempt that enabled butyrate production from a non-butyrate producing commensal B. thetaiotaomicron. The study also highlights that B. thetaiotaomicron can serve as an effective strain for live microbial therapeutics in human.

Bacteroides thetaiotaomicron代表了人类肠道微生物组的主要共生体，越来越多地被视为微生物治疗的有希望的候选菌株。在这里，我们设计了B. thetaiotaomicron，用于异源生产非天然丁酸盐作为治疗相关浓度的概念验证生化。由于B. thetaiotaomicron不是丁酸盐的天然生产者，我们在菌株中异源表达了丁酸盐生物合成途径，这导致在富培养基中产生终浓度为 12 mg/L 的丁酸盐。通过由B. thetaiotaomicron的扩展基因组规模代谢模型 (GEM) 指导的一轮代谢工程，实现了丁酸盐生产的进一步优化。这扩展模型的计算机模拟敲除模拟表明pta和ldhD是提高丁酸盐产量的有效敲除目标。pta - ldhD双敲除突变体中丁酸盐的最大效价和比生产力分别增加了近3.4倍和4.8倍。据我们所知，这是第一次工程尝试，能够从不产生丁酸盐的共生B. thetaiotaomicron 中生产丁酸盐。该研究还强调B. thetaiotaomicron可以作为人类活微生物疗法的有效菌株。