Applied Microbiology and Biotechnology, Vol.104, No.6, 2545-2559, 2020
Pathway construction and metabolic engineering for fermentative production of beta-alanine in Escherichia coli
beta-Alanine is a naturally occurring beta-amino acid that has been widely applied in the life and health field. Although microbial fermentation is a promising method for industrial production of beta-alanine, an efficient microbial cell factory is still lacking. In this study, a new metabolically engineered Escherichia coli strain for beta-alanine production was developed through a series of introduction, deletion, and overexpression of genes involved in its biosynthesis pathway. First, the L-aspartate a-decarboxylase gene, BtADC, from Bacillus tequilensis, with higher catalytic activity to produce beta-alanine from aspartate, was constitutively expressed in E. coli, leading to an increased production of beta-alanine up to 2.76 g/L. Second, three native aspartate kinase genes, akI, akII, and akIII, were knocked out to promote the production of beta-alanine to a higher concentration of 4.43 g/L by preventing from bypass loss of aspartate. To increase the amount of aspartate, the native AspC gene was replaced with PaeAspDH, a L-aspartate dehydrogenase gene from Pseudomonas aeruginosa, accompanied with the overexpression of the native AspA gene, to further improve the production level of beta-alanine to 9.27 g/L. Last, increased biosynthesis of oxaloacetic acid (OAA) was achieved by a combination of overexpression of the native PPC, introduction of CgPC, a pyruvate decarboxylase from Corynebacterium glutamicum, and deletion of ldhA, pflB, pta, and adhE in E. coli, to further enhance the production of beta-alanine. Finally, the engineered E. coli strain produced 43.12 g/L beta-alanine in fed-batch fermentation. Our study will lay a solid foundation for the promising application of beta-alanine in the life and health field.