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Enzyme mining and D-tagatose synthesis using a dual-enzyme redox system
Yin Fang1, Jiangbo Li2, Na Li1
1College of Food Science and Light Industry, Nanjing Tech University, Nanjing, 211816, China.
Biochemical and Biophysical Research Communications
|July 8, 2025
Summary
Researchers developed an efficient D-tagatose production method using a dual-enzyme system in E. coli. This advances the synthesis of the low-calorie sweetener D-tagatose, offering a promising alternative for managing diabetes and obesity.
Area of Science:
- Biotechnology
- Enzyme Engineering
- Metabolic Engineering
Background:
- Global rise in diabetes and obesity necessitates healthier sweetener alternatives.
- D-tagatose, a low-calorie sweetener, shows therapeutic potential but faces production challenges.
- Current D-tagatose synthesis methods using L-arabinose isomerase or redox enzymes have limitations in conversion rates and purification.
Purpose of the Study:
- To develop a highly efficient D-tagatose production method using a novel dual-enzyme redox system.
- To overcome the limitations of thermodynamic equilibrium and low enzyme activity in previous D-tagatose synthesis routes.
- To engineer an E. coli cell factory for the optimized biosynthesis of D-tagatose.
Main Methods:
- Enzyme screening identified high-activity xylose reductase from Meyerozyma guilliermondii (MgXR) and galactitol dehydrogenase from Rhizobium meliloti (RmGDH).
- Co-expression of MgXR and RmGDH in E. coli BL21(DE3) for D-galactose conversion.
- Implementation of a NADPH regeneration system to enhance reaction efficiency.
- Fed-batch fermentation of engineered E. coli MgRm-trcf strain using lactose as substrate.
Main Results:
- MgXR exhibited an activity of 42.2 ± 1.1 U/mg, and RmGDH showed an activity of 10.2 ± 0.8 U/mg.
- Co-expression yielded an initial D-tagatose titer of 15.48 g/L from D-galactose.
- The addition of a NADPH regeneration system increased the D-tagatose titer to 21.3 g/L.
- Fed-batch fermentation produced a record 34.7 g/L D-tagatose with a productivity of 0.482 g/L·h.
Conclusions:
- The engineered E. coli cell factory with a dual-enzyme redox system provides a highly efficient route for D-tagatose synthesis.
- This study demonstrates a significant advancement in the biotechnological production of D-tagatose, surpassing previous yields.
- The developed method offers a sustainable and scalable approach for producing D-tagatose, a valuable sweetener for health-conscious markets.
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