Engineering Compartmentalization and Cofactor Regulation for 10-Hydroxy-2-decenoic Acid Biosynthesis in Saccharomyces...

Area of Science:

• Biotechnology
• Metabolic Engineering
• Synthetic Biology

Background:

• 10-Hydroxy-2-decenoic acid (10-HDA) is a valuable royal jelly component with pharmacological properties.
• Previous biosynthesis of 10-HDA was limited to Escherichia coli, posing challenges for biomedical use due to endotoxin concerns.

Purpose of the Study:

• To engineer a Generally Recognized As Safe (GRAS) yeast Saccharomyces cerevisiae platform for 10-HDA production.
• To overcome the limitations of E. coli-based biosynthesis for 10-HDA.

Main Methods:

• Rewiring of the beta-oxidation pathway in S. cerevisiae.
• Mitochondrial compartmentalization with chaperone-assisted P450 folding.
• Enhancement of NADPH supply.
• Fed-batch fermentation using ethyl decanoate to mitigate substrate toxicity.

Main Results:

• Achieved an initial yield of 40.50 mg/L of 10-HDA using decanoic acid as substrate.
• Increased production 7.5-fold via fed-batch fermentation with ethyl decanoate, reaching 298.6 mg/L.
• Established the highest titer of 10-HDA reported in yeast to date.

Conclusions:

• Successfully developed a GRAS-compliant yeast platform for safe and efficient 10-HDA production.
• Demonstrated a novel strategy for functional expression of prokaryotic P450 enzymes in yeast.
• This platform facilitates broader biomedical applications of 10-HDA.