As an important dicarboxylic acid, malonic acid is widely used in many fields. However, due to the low yield of malonic acid synthesized by biological method, it is not suitable for industrial production currently. To improve the biosynthesis of malonic acid, a recombinant strain BL21(PPP) was constructed to synthesize malonate by overexpressing six genes of ppc, aspA, panD, sdhC, pa0132 and yneI by using Escherichia coli BL21(DE3) as the chassis cell. The fermentation conditions of this strain were optimized in shake flask, and it was found that the strain could produce 0.48 g/L malonic acid, which was 1-fold higher than before optimization. The effects of additional biotin and fumaric acid on malonic acid production were also investigated, and it was found that the production of malonic acid was increased by 32.59% compared with the control group. In addition, the strain could produce 1.2 g/L malonic acid at a final concentration of 8 g/L fumaric acid. Finally, in a 5 L fermenter, the production of malonic acid was increased to 12.42 g/L. In this study, the biosynthesis of malonic acid in E. coli BL21(DE3) was achieved, and the yield of malonic acid was further improved by the optimization of fermentation, which laid a foundation for further improvement of malonic acid production.
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