[Efficient biosynthesis of _D-mannitol by coordinated expression of a two-enzyme cascade]

Area of Science:

• Biotechnology
• Enzyme Engineering
• Metabolic Engineering

Background:

• D-mannitol is a valuable precursor in pharmaceuticals, but current synthesis methods are costly.
• Efficient and scalable production of D-mannitol is crucial for its wider application.

Purpose of the Study:

• To develop a cost-effective and efficient enzymatic method for D-mannitol synthesis.
• To engineer a recombinant E. coli strain for cascade biocatalysis of D-fructose to D-mannitol.

Main Methods:

• Screening and co-expression of mannitol dehydrogenase (LpGDH) and glucose dehydrogenase (BaGDH) in E. coli BL21(DE3).
• Engineering a recombinant strain (E. coli BL21/pETDuet-Lpmdh-Bagdh-Bagdh) to enhance NADH regeneration.
• Optimization of whole-cell biotransformation conditions (temperature, pH, substrate concentration).

Main Results:

• Achieved a 59.7% conversion rate of D-fructose to D-mannitol using a two-enzyme cascade.
• Optimized whole-cell biotransformation yielded 81.9 g/L D-mannitol with an 81.9% molar conversion rate.
• Demonstrated enhanced NADH regeneration by increasing BaGDH copy number.

Conclusions:

• Developed a green and efficient whole-cell biotransformation method for D-mannitol production.
• The engineered E. coli strain and optimized process are suitable for large-scale D-mannitol manufacturing.
• This approach holds significance for the production of other sugar alcohols.