丙二酸作为一种重要的二元羧酸,广泛应用于多种领域。但目前由于生物法合成丙二酸产量较低,不适合进行工业化生产。为提高丙二酸的生物法合成能力,该文以大肠杆菌BL21(DE3)为底盘细胞,通过过量表达ppc、aspA、panD、sdhC、pa0132及yneI基因,构建了丙二酸生物合成工程菌BL21(PPP)。在摇瓶条件下,对该菌株的发酵条件进行了优化,丙二酸合成量达0.48 g/L,较优化前提高了1倍。同时,该文探究了外源添加生物素及富马酸对丙二酸合成的影响,在添加75 μg/L生物素的发酵条件下,丙二酸的产量较对照组提高了32.59%;在添加终质量浓度为8 g/L的富马酸的条件下,该菌株可合成1.2 g/L丙二酸。在5 L发酵罐中,该菌株最高可以合成12.42 g/L的丙二酸。该文实现了丙二酸在大肠杆菌BL21(DE3)中的生物合成,并通过对发酵条件的优化进一步提高了丙二酸的产量,为更进一步提高丙二酸产量奠定了基础。
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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