Engineering Dirhodium Artificial Metalloenzymes for Diazo Coupling Cascade Reactions

Artificial metalloenzymes (ArMs) are commonly used to control the stereoselectivity of catalytic reactions, but controlling chemoselectivity remains challenging. In this study, we engineer a dirhodium ArM to catalyze diazo cross‐coupling to form an alkene that, in a one‐pot cascade reaction, is reduced to an alkane with high enantioselectivity (typically >99 % ee) by an alkene reductase. The numerous protein and small molecule components required for the cascade reaction had minimal effect on ArM catalysis. Directed evolution of the ArM led to improved yields and E/Z selectivities for a variety of substrates, which translated to cascade reaction yields. MD simulations of ArM variants were used to understand the structural role of the cofactor on ArM conformational dynamics. These results highlight the ability of ArMs to control both catalyst stereoselectivity and chemoselectivity to enable reactions in complex media that would otherwise lead to undesired side reactions. Directed evolution was used to improve dirhodium metalloenzyme (ArM)‐catalyzed diazo cross‐coupling yield and selectivity. The ArM was then used in cascade reactions where the alkene product was reduced by an ene reductase, with the ArM scaffold controlling chemoselectivity in the reaction mixture. MD simulations show that the cofactor modulates the domain dynamics and favors a more closed, hydrophobic pocket conducive to selective catalysis.