Improvement of acetate tolerance of Escherichia coli by introducing the PHB mobilization pathway.

Poly-β-hydroxybutyrate (PHB) mobilization has been recognized as an effective measure in bacteria for host survival under stressful conditions. Here, we report that PHB mobilization is also involved in improving Escherichia coli resistance to acetic acid stress. Under 0.06% (vol/vol) acetic acid stress, the cell viability of strain M5 with PHB mobilization reached 52.8%, significantly higher than the 25.2% observed for strain M1 without PHB mobilization. Comparison of transcriptome data between M1 and M5 strains identified genes associated with membrane formation that participate in acetic acid tolerance. As a result, the membrane integrity of the M5 strain was significantly increased by 31.5% compared to strain M1. According to physiological membrane analysis, strain M5 showed a 35.3% increase in the ratio of cyclic to unsaturated fatty acids compared to strain M1 under 0.06% (vol/vol) acetic acid. These results indicate that PHB mobilization can regulate membrane components, consequently enhancing cell tolerance to acetic acid. Furthermore, acetate serves as a cost-effective alternative carbon source for microbial cultivation. Then, the succinate-producing strain M8 containing PHB mobilization was used to produce succinate and PHB from sodium acetate. Interestingly, PHB mobilization improved sodium acetate tolerance and utilization in E. coli, while enhancing succinate and PHB production. Finally, strain M8 can accumulate 23.93 g/L succinate and 7.21 g/L PHB using sodium acetate under fed-batch fermentation. In conclusion, this work reveals the role of PHB mobilization under acetate stress and provides a basis for acetate utilization.IMPORTANCEThis study investigated the underlying mechanism through which PHB mobilization enhances Escherichia coli tolerance to acetic acid stress. PHB mobilization improved E. coli tolerance to acetic acid, leading to enhanced cell viability. The transcriptome results indicated that PHB mobilization mainly alters the expression of membrane-associated genes, such as gene Bhsa (encoding outer membrane protein), leading to increased resistance to acetic acid. The membrane physiological analysis indicated that PHB mobilization plays a critical role in membrane integrity, fluidity, and lipid components under acetic acid stress. Moreover, we proposed a novel approach for the co-synthesis of succinate and PHB in recombinant E. coli from sodium acetate. The succinate-producing strain M8 harboring PHB mobilization can efficiently co-produce succinate and PHB, exhibiting better cell growth and sodium acetate utilization compared to the control strain without PHB mobilization. These findings indicate that PHB mobilization has implications for developing robust E. coli and their biosynthesis applications.