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Glutaric acid production by systems metabolic engineering of an l-lysine–overproducing Corynebacterium glutamicum

Significance Glutaric acid is an important dicarboxylic acid used in the synthesis of commercial polymers. Here we report metabolic engineering of an l-lysine–overproducing Corynebacterium glutamicum strain for the high-level production of glutaric acid. The glutaric acid biosynthesis pathway was established in C. glutamicum by optimally expressing the genes encoding key enzymes from Pseudomonas putida and C. glutamicum for the efficient conversion of l-lysine to glutaric acid. Further metabolic engineering and overexpression of a newly discovered glutaric acid exporter gene resulted in the production of 105.3 g/L of glutaric acid without byproducts from glucose. The metabolic engineering strategies described here will be useful for developing microbial strains for the bio-based production of other chemicals in addition to glutaric acid. There is increasing industrial demand for five-carbon platform chemicals, particularly glutaric acid, a widely used building block chemical for the synthesis of polyesters and polyamides. Here we report the development of an efficient glutaric acid microbial producer by systems metabolic engineering of an l-lysine–overproducing Corynebacterium glutamicum BE strain. Based on our previous study, an optimal synthetic metabolic pathway comprising Pseudomonas putida l-lysine monooxygenase (davB) and 5-aminovaleramide amidohydrolase (davA) genes and C. glutamicum 4-aminobutyrate aminotransferase (gabT) and succinate-semialdehyde dehydrogenase (gabD) genes, was introduced into the C. glutamicum BE strain. Through system-wide analyses including genome-scale metabolic simulation, comparative transcriptome analysis, and flux response analysis, 11 target genes to be manipulated were identified and expressed at desired levels to increase the supply of direct precursor l-lysine and reduce precursor loss. A glutaric acid exporter encoded by ynfM was discovered and overexpressed to further enhance glutaric acid production. Fermentation conditions, including oxygen transfer rate, batch-phase glucose level, and nutrient feeding strategy, were optimized for the efficient production of glutaric acid. Fed-batch culture of the final engineered strain produced 105.3 g/L of glutaric acid in 69 h without any byproduct. The strategies of metabolic engineering and fermentation optimization described here will be useful for developing engineered microorganisms for the high-level bio-based production of other chemicals of interest to industry.