A growth-coupled progesterone-responsive biosensor for high-throughput microfluidic screening in Saccharomyces...

Area of Science:

• Synthetic biology
• Biotechnology
• Microbial biotransformation

Background:

• Poor aqueous solubility of steroid precursors like pregnenolone and progesterone hinders microbial biotransformation and strain screening.
• This limitation presents a bottleneck for improving yeast strains for industrial steroid production.

Purpose of the Study:

• To develop a high-throughput screening platform for enhancing steroid production in yeast.
• To overcome the solubility limitations of steroid precursors using a novel biosensor and cyclodextrin system.

Main Methods:

• Developed a growth-coupled progesterone-responsive biosensor in Saccharomyces cerevisiae, integrated with hydroxypropyl-β-cyclodextrin (HP-β-CD).
• Utilized atmospheric and room temperature plasma (ARTP) mutagenesis for strain improvement.
• Employed microfluidic droplet encapsulation for high-producer separation.

Main Results:

• Identified five yeast variants with 2.0-3.37-fold higher progesterone production compared to wild-type strains.
• Demonstrated the platform's selectivity, robustness, and scalability in separating high-producing cells.
• Validated the improved conversion of pregnenolone to progesterone in selected variants.

Conclusions:

• The developed synthetic biology platform offers a practical, modular, and high-throughput strategy for screening poorly water-soluble steroid-producing yeast.
• The system is adaptable for other bioactive molecules and shows potential for enzyme evolution and broader biotechnological applications.