Efficient L-valine production using systematically metabolic engineered Klebsiella oxytoca

Area of Science:

• Metabolic Engineering and Synthetic Biology
• Microbial Fermentation
• Biochemical Production

Background:

• L-Valine, an essential branched-chain amino acid, has diverse industrial applications.
• Current microbial production methods face challenges in yield and efficiency.
• Klebsiella oxytoca is a known 2,3-butanediol producer, requiring metabolic redirection for L-valine synthesis.

Purpose of the Study:

• To engineer Klebsiella oxytoca for efficient L-valine biosynthesis.
• To redirect metabolic flux from 2,3-butanediol to L-valine production.
• To optimize fermentation conditions for high-titer L-valine.

Main Methods:

• Metabolic flux analysis and pathway engineering in Klebsiella oxytoca.
• Introduction of exogenous L-valine biosynthesis genes.
• Blocking of endogenous 2,3-butanediol pathway at the α-acetolactate precursor.
• Enhancement of L-valine efflux and pyruvate polymerization.
• Construction of a plasmid-free engineered strain (K. oxytoca VKO-9).

Main Results:

• Engineered K. oxytoca VKO-9 produced 122 g/L L-valine with a yield of 0.587 g/g in 56 hours via fed-batch fermentation.
• Repeated fed-batch fermentation achieved average L-valine concentration of 81.3 g/L, yield of 0.599 g/g, and productivity of 3.39 g/L/h.
• Metabolic redirection successfully shifted carbon flux towards L-valine synthesis.

Conclusions:

• The developed metabolic engineering strategy is effective for high-titer L-valine production using Klebsiella oxytoca.
• Optimized fermentation processes, including repeated fed-batch, enhance productivity and manage product precipitation.
• This approach offers a promising platform for sustainable microbial production of L-valine.