A Cellular Coupling Strategy Leveraging Pathway Modularization and Cofactor Regeneration for the Biosynthesis of...

Area of Science:

• Biotechnology and Metabolic Engineering
• Synthetic Biology
• Carbohydrate Chemistry

Background:

• 3 -Sialyllactose (3′-SL) is a crucial sialylated human milk oligosaccharide vital for infant development and physiological functions.
• Existing methods for 3′-SL synthesis are often complex and may not be scalable for industrial production.

Purpose of the Study:

• To establish a synergistic whole-cell catalysis system for the efficient biosynthesis of 3′-SL.
• To optimize the engineered microbial strains and fermentation conditions for maximizing 3′-SL production.

Main Methods:

• Engineered a multi-module whole-cell biocatalyst using genes from Escherichia coli and Saccharomyces cerevisiae.
• Utilized pyruvate, N-acetylglucosamine, lactose, and cytidine-5 -monophosphate as substrates for synergistic synthesis.
• Optimized induction conditions and substrate dosage in engineered E. coli JM109(DE3) for whole-cell catalysis.

Main Results:

• Achieved a 3′-SL titer of 71.62 g/L after optimization of induction and substrate dosage.
• Successfully scaled up the process in a 5 L bioreactor, reaching a maximum 3′-SL titer of 78.03 g/L.
• Demonstrated the first report of synergistic 3′-SL synthesis using three modules from E. coli and S. cerevisiae.

Conclusions:

• The developed whole-cell catalysis system provides an efficient and scalable route for 3′-SL production.
• This biosynthetic approach holds significant potential for producing functional oligosaccharides for infant nutrition and health.
• The study highlights the power of combining enzymes from different microbial hosts for complex molecule synthesis.