Engineering cofactor flexibility enhanced 2,3-butanediol production in Escherichia coli

Area of Science:

• Biotechnology
• Metabolic Engineering
• Synthetic Biology

Background:

• 2,3-butanediol (2,3-BD) production relies on enzymes requiring nicotinamide adenine dinucleotide (NADH) or its phosphate form (NADPH) as electron donors.
• Biosynthesis efficiency is limited by enzyme specificity and dynamic cofactor availability.

Purpose of the Study:

• To engineer cofactor flexibility for enhanced 2,3-BD production.
• To improve stereo-specificity and yield of 2,3-BD biosynthesis.

Main Methods:

• Simultaneous overexpression of an NADH-dependent 2,3-BD dehydrogenase (KpBudC) and an NADPH-specific 2,3-BD dehydrogenase (CbAdh).
• Optimization of medium composition and fermentation conditions.

Main Results:

• Co-expression enabled versatile cofactor utilization for 2,3-BD synthesis.
• Achieved 92 g/L of 2,3-BD in 56 hours with 90% stereo-purity for the (R,R)-isoform.
• Reached 85% of the maximum theoretical yield without acetoin accumulation.

Conclusions:

• Cofactor flexibility is a key design principle for improving bio-based chemical production involving redox reactions.
• Engineered strains offer a robust platform for efficient and stereo-specific 2,3-BD manufacturing.