Rewiring P450s for efficient biocatalysis

This project combines novel P450 catalysts with cutting edge enzyme engineering techniques to selectively hydroxylate substrate molecules. P450s have enormous potential for the sustainable manufacture of both fine chemicals (fragrances, flavourings, pharmaceuticals) and commodity chemicals (fuels, solvents, agrochemicals) but are limited by key electron transfer steps in their mechanisms. In this project, new fusion proteins will be engineered, in which electron transfer pathways between haem and reductase domains are optimised to maximise catalytic efficiency. The new "re-wired" P450s will then be investigated by state of the art electron paramagnetic resonance (EPR) spectroscopy. Here, two spin active species (either native to the protein, or added using nitroxide spin labels) will be incorporated and used to obtain nanoscale distance measurements between these labels situated in the reductase and haem domains using EPR under catalytically relevant conditions. Further study by computational techniques will rationalise the findings and thereby provide a full understanding of how P450s can be rewired to maximise their utility for industrial applications.