Metabolic engineering of Corynebacterium glutamicum CGY-PG-304 for promoting gamma-aminobutyric acid production

Abstract

Gamma-aminobutyric acid is a versatile and non-protein amino acid that plays a significant role in medicine, food, and cosmetics. The synthesis of gamma-aminobutyric acid is restricted by complex metabolic mechanisms and suboptimal fermentation conditions. Previously, we had constructed the Corynebacterium glutamicum strain CGY-PG-304 which could efficiently produce gamma-aminobutyric acid. In this study, we promoted gamma-aminobutyric acid production in CGY-PG-304 by enhancing the carbon flow in the TCA cycle, streamlining the mycolic acid layer of the cell wall, and optimizing the fermentation conditions. First, the genes sucCD encoding succinyl coenzyme A synthase, the gene cmrA encoding the ketoacyl reductase, and the gene treY encoding maltooligosaccharyl trehalose synthase were deleted in CGY-PG-304 individually or in combination. The yield of gamma-aminobutyric acid was increased in all the resulting strains among which CGW003 was the best. Next, the gene acnA encoding cis-aconitase or the gltS encoding sodium-coupled glutamate secondary uptake system were overexpressed in CGW003 using plasmid, and the former produced more gamma-aminobutyric acid than the latter. Therefore, the promoter of the chromosomal gene acnA in CGW003 was replaced by the strong promoter PtacM, resulting in the final strain CGW005. CGW005 could produce 112.03 g/L of gamma-aminobutyric acid with a yield of 0.34 g/g of glucose by fed-batch fermentation.

Funding

National Key Research and Development Program of China,(2021YFC2100900)