Metabolic Engineering of Escherichia coli for the Production of 5-Aminolevulinic Acid Based on Combined Metabolic Pathway Modification and Reporter-Guided Mutant Selection (RGMS)

Abstract

Abstract Background 5-Aminolevulinic acid (ALA) recently received much attention due to its potential application in many fields such as medicine, nutrition and agriculture. Metabolic engineering is an efficient strategy to improve microbial production of 5-ALA. Results In this study, an ALA production strain of Escherichia coli was constructed by rational metabolic engineering and stepwise improvement. A metabolic strategy to produce ALA directly from glucose in this recombinant E. coli via both C4 and C5 pathways was applied herein. The expression of a modified hemARS gene and rational metabolic engineering by gene knockouts significantly improved ALA production from 765.9 to 2056.1 mg/L. Next, we tried to improve ALA production by RGMS-directed evolution of eamA gene. After RGMS, the ALA yield of strain A2-ASK reached 2,471.3 mg/L in flask. Then we aimed to improve the oxidation resistance of cells by overexpressing sodB and katE genes and ALA yield reached 2703.8 mg/L. A final attempt is to replace the promoter of hemB gene in genome to inhibit the hemB gene. After 24 h cultivation, a high ALA yield of 19.02 g/L was achieved by 108-ASK in a 5 L fermenter. Conclusions These results suggested that an industrially competitive strain can be efficiently developed by metabolic engineering based on combined rational modification and optimization of gene expression.