Design and Evolution of Photoenzymes for Triplet Energy Transfer Catalysis

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The ability to program new modes of catalysis into enzymes would have profound impacts across chemistry and biotechnology, delivering sustainable biocatalytic processes to address societal needs. Genetic code expansion technology opens up exciting new opportunities in biocatalyst design and engineering, by allowing site-selective introduction of new functional elements into proteins as non-canonical amino acid side chains. In TRIPase, I will exploit an expanded genetic code to develop efficient, selective and programmable photoenzymes that operate via triplet energy transfer processes, a versatile mode of reactivity in organic synthesis that is currently not accessible to biocatalysis. These photoenzymes will contain functional elements needed to harness light energy and promote valuable chemical reactions including [2+2]cycloadditions, electrocyclizations, rearrangements and deracemizations. My approach will employ engineered translation components to introduce organic photosensitizers into protein active sites. Protein cavities offer attractive and versatile chiral environments for mediating enantioselective photochemistry, where substrates and key catalytic elements can be accurately positioned within a single pocket. Since the photosensitizers are genetically encoded, active photoenzymes can be optimized via directed evolution to enhance catalytic efficiency and quantum yields, or to impart new functions that are challenging to achieve with small molecule photocatalysts. Structural and biochemical analysis of engineered photoenzymes will shed light on the active site features and mechanistic strategies responsible for enhanced photocatalysis to guide future biocatalyst design. Overall, the platform technology developed in TRIPase will open the door to a wealth of new excited-state chemistry in proteins and in doing so will underpin the development of a new generation of evolvable photocatalysts with efficiencies and specificities akin to natural enzymes.