A Toolbox for Diverse Oxyfunctionalisation of Monoterpenes

Area of Science:

• Synthetic biology
• Biocatalysis
• Enzyme engineering

Background:

• Effective chemical biosynthesis requires extensive libraries of well-characterized enzymes.
• Synthetic biology methodologies can be leveraged to accelerate the creation of such enzyme libraries.

Purpose of the Study:

• To develop a scalable pipeline for generating and characterizing enzyme libraries using synthetic biology.
• To create a cytochrome P450 enzyme collection for the oxyfunctionalization of terpene scaffolds.

Main Methods:

• Designed and constructed a cytochrome P450 enzyme collection.
• Employed a custom untargeted gas chromatography-mass spectrometry (GC/MS)-metabolomics approach for characterization.
• Applied synthetic biology principles to bootstrap library creation.

Main Results:

• Successfully generated and characterized a comprehensive enzyme library.
• Demonstrated a wide range of activities and selectivities for oxyfunctionalizing monoterpenes.
• Established a novel resource for accessing terpenoid chemical diversity.

Conclusions:

• The developed pipeline offers a rapid and scalable method for enzyme library creation.
• The cytochrome P450 enzyme collection facilitates controlled oxyfunctionalization of terpenes.
• This resource enables pathway engineering for producing diverse oxygenated terpenoids for applications like drug discovery.