Engineering Klebsiella oxytoca for enhanced 2,3-butanediol production from gluconate

Abstract

Historically, microbial production of 2,3-BD has relied predominantly on glucose as the substrate. However, its fermentation necessitates microaerobic conditions due to inherent redox imbalances. This limitation complicates process control and can lead to undesirable late-stage consumption of 2,3-BD. This study explored the use of gluconate as an alternative substrate for 2,3-butanediol (2,3-BD) production in Klebsiella oxytoca . This study reveals that the conversion of gluconate to 2,3-BD is redox balanced, allowing for a streamlined anaerobic fermentation process, which is industrially advantageous and circumvents the late-stage 2,3-BD consumption observed in glucose microaerobic fermentation. The wild-type K. oxytoca DSM 4798 strain achieved a 2,3-BD yield of 47% and 54% from gluconate and glucose, respectively, under anaerobic conditions. The MJ09 strain, engineered through targeted deletions ( ∆ldhA ∆ackA-pta ∆frdABCD ∆adhE ), demonstrated a significant enhancement, achieving a 2,3-BD yield of 90% from gluconate under anaerobic conditions. However, the same strain lost the ability to grow on glucose. These results highlight gluconate as a promising substrate for anaerobic 2,3-BD production, offering a redox-balanced pathway that simplifies fermentation. Deleting competing pathways successfully improved the 2,3-butanediol yields.