My research is focused on redox enzymes, from protein recognition and kinetics of electron transfer processes to their biotechnological applications in green and sustainable synthesis. My group has a long-standing interest in the cytochrome P450 family of monooxygenases. These enzymes catalyse the direct oxidation of unactivated C–H bonds in complex organic molecules using oxygen in air as the oxidant. The primary catalytic activity is conversion to the alcohol derivative but unusual reactions such as C–C bond formation have also been discovered. Our research ranges from discovering and characterising the substrate range of new P450 enzymes to designing and evolving these enzymes for scalable late-stage C–H activation. Recent targets include synthetic biology approaches to aroma and flavour production, oxidative diversification of core structures of agrochemical and pharmaceutical intermediates, and chemoenzymatic cascades for more efficient synthesis routes.
Discovering and engineering P450 enzymes
Metabolically diverse bacteria often contain oxidative enzymes such as cytochrome P450s to catalyse the initial oxidation step to activate chemically inert compounds in metabolic and catabolic processes, and for the detoxification of xenobiotics. As such, they are rich sources of enzymes with unusual and often broad substrate range. We are interested in the cloning, heterologous expression and activity profile of new enzymes for developing new processes.
Aroma and flavour fine chemicals
Some of the most desirable aromas and flavours are oxygenated natural hydrocarbons. The biosynthesis of these alcohols, aldehydes, ketones, esters and lactones typically involve an oxygenation step catalysed by a P450 enzyme. One of our main research programmes is the engineering and evolution of the enzyme P450BM3 to catalyse this final step in the synthesis of a wide range of aroma and flavour compounds. The conversion of valencene from orange essential oil to nootkatone, the grapefruit flavour, is a commercial process. Other targets include the rose ketones, fruity lactones and wood essential oil aroma compounds. The high value of many of these compounds stems from the low natural abundance of both the precursor and the final product. My group is developing synthetic biology systems for the total biosynthesis of these compounds in a single organism.
Late-stage C–H activation of drugs and drug-like compounds
We have developed a library of over 400 P450 enzyme variants with different combinations of amino acid substitutions to alter the size, shape and polarity of the substrate binding pocket. Subsets of this collection of variants are screened for the oxidation of organic compounds to explore the activity and product diversity. The trends are then used to design and evolve the variants to provide high selectivity for the greatest range of C–H bond oxidation products. This library approach is applied to late-stage C–H activation in synthesis, whereby the carbocyclic core is assembled first, and then the key intermediates are selectively oxidised, potentially providing truncated and more efficient synthetic pathways. This approach has been applied to natural product synthesis (with Jeremy Robertson).
Selective oxidation to a wide range of alcohol and carbonyl derivatives has important applications in diversity-oriented synthesis (DOS). Stereoselective oxidation to the alcohol group introduces a versatile synthetic handle for functional group elaboration. DOS is particularly relevant in drug discovery and synthesis, as diversely functionalised compounds are less lipophilic and may show stronger and more specific binding to drug targets. We have demonstrated oxidative diversification of privileged drug core motifs such as steroids, tetrahydroquinolines and heterocyclic compounds.