Systems metabolic engineering of Corynebacterium glutamicum towards improved lysine production

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L-lysine, an essential amino acid primarily used as a supplement in animal feed, has a global market of approximately 1,000,000 t/a, making it a significant fermentation product. The gram-positive soil bacterium Corynebacterium glutamicum is the main organism used for its production. This work focuses on the rational strain engineering of lysine production through a systems-oriented approach. Detailed investigations revealed that lysine biosynthesis, NADPH metabolism, the TCA cycle, and precursor supply are crucial pathways. A central method employed was 13C metabolic flux analysis, providing insights into cellular physiology. The study extended an existing model to enhance the number of estimable flux parameters. For strain optimization, targeted modulation of enzyme activities was essential to adjust carbon conversion in selected pathways. Techniques included promoter exchange, gene deletion, and a novel start codon exchange method for permanent enzyme activity modulation. Genetically defined wild-type lysine producers expressing a feedback-deregulated variant of aspartate kinase served as host strains. Key modifications included overexpression of diaminopimelate dehydrogenase, down-regulation of the TCA cycle through isocitrate dehydrogenase attenuation, and NADPH metabolism engineering. Implementing these targets in a single strain resulted in a lysine hyper-producer with impressive production capabilities, achieving a lys