Alternate Heme Ligation Steers Activity and Selectivity in Engineered Cytochrome P450-Catalyzed Carbene-Transfer Reactions

We report a biocatalytic platform of engineered cytochrome P450 enzymes to carry out carbene-transfer reactions using a lactone-based carbene precursor. By simply altering the heme-ligating residue, we obtained two enzymes that catalyze olefin cyclopropanation (Ser) or S–H bond insertion (Cys). Both enzymes exhibit high catalytic efficiency and stereo­selectivity, thus enabling facile access to structurally diverse spiro[2.4]­lactones and α-thio-γ-lactones. Computational studies revealed the mechanism of carbene S–H insertion and explain how the axial ligand controls reactivity and selectivity. This work expands the catalytic repertoire of heme­proteins and offers insights into how these enzymes can be tuned for new chemistry.