Structure-Guided Design of Artificial Transcription Factor for a Progesterone Biosensor

Area of Science:

• Synthetic biology
• Molecular biology
• Biotechnology

Background:

• Transcription factors (TFs) are crucial for gene regulation in synthetic biology.
• Eukaryotic TFs are rarely utilized in prokaryotic systems for gene control.
• Developing novel TFs for prokaryotic systems is essential for advancing synthetic biology.

Purpose of the Study:

• To design and construct an artificial TF responsive to progesterone.
• To create a whole-cell biosensor for progesterone detection in E. coli.
• To optimize the biosensor's performance for potential clinical applications.

Main Methods:

• Artificial TF ProB designed using MD simulations, incorporating QF_DBD, ligand-binding domain, linker, and QF_AD.
• ProB regulated a synthetic promoter QT controlling GFP expression in E. coli.
• Bacterial enrichment strategy employed for biosensor optimization.

Main Results:

• The ProB TF successfully controlled gene transcription in E. coli.
• The whole-cell progesterone biosensor exhibited high sensitivity (LOD 0.15 μg/L, EC₅₀ 26.58 μg/L).
• Optimized biosensor demonstrated rapid detection (56.4 min) within a working range (0.15-40 μg/L) and preferred temperature (20-30 °C).

Conclusions:

• An artificial TF and a progesterone biosensor were successfully developed for prokaryotic systems.
• The engineered biosensor meets clinical application requirements for progesterone detection.
• This study provides valuable insights into designing non-natural TFs and artificial gene circuits in prokaryotes.