Synthetic translational coupling system for accurate and predictable polycistronic gene expression control in...

Area of Science:

• Synthetic Biology
• Molecular Biology
• Metabolic Engineering

Background:

• Inadequate control of polycistronic gene expression leads to suboptimal metabolic flux and imbalanced protein assembly in bacteria.
• Precise and predictable genetic elements are crucial for addressing these challenges in microbial systems.

Purpose of the Study:

• To devise a synthetic biopart based on translational coupling to control polycistronic gene expression.
• To enable precise and predictable control over gene expression ratios within polycistronic constructs.

Main Methods:

• Engineered synthetic bioparts utilizing translational coupling to link genes on a polycistronic mRNA.
• Modified Shine-Dalgarno sequences within the bioparts to tune relative gene expression.
• Created 41 distinct bioparts with tunable expression ratios ranging from 0.03 to 0.92.

Main Results:

• Achieved precise control over polycistronic gene expression ratios, independent of coding sequences or transcription rates.
• Demonstrated up to a 7.6-fold increase in the production of valuable biochemicals, including 3-hydroxypropionic acid, poly(3-hydroxybutyrate), and lycopene.
• Successfully applied the bioparts for pathway enzyme gene expression in a polycistronic manner.

Conclusions:

• Developed novel genetic regulatory modules for precise and predictable polycistronic gene expression.
• Facilitated efficient protein assembly, biosynthetic gene cluster expression, and pathway optimization in bacteria.
• Enabled enhanced production of valuable biochemicals through improved genetic control.