A Toolbox for Diverse Oxyfunctionalisation of Monoterpenes

The successful implementation of synthetic biology for chemicals biosynthesis relies on the availability of large libraries of well-characterized enzymatic building blocks. Here we present a scalable pipeline that applies the methodology of synthetic biology itself to bootstrap the creation of such a library. By designing and building a cytochrome P450 enzyme collection and testing it in a custom-made untargeted GC/MS-metabolomics-based approach, we were able to rapidly create and characterize a comprehensive enzyme library for the controlled oxyfunctionalisation of terpene scaffolds with a wide range of activities and selectivities towards several monoterpenes. This novel resource can now be used to access the extensive chemical diversity of terpenoids by pathway engineering and the assembly of biocatalytic cascades to subsequently produce libraries of oxygenated terpenoids and their derivatives for diverse applications, including drug discovery.