Transition metal–hydrides have been widely exploited in catalysis for the hydrofunctionalization of unsaturated moieties, including carbonyls, alkenes and alkynes1. To complement heterolytic metal–hydride bond cleavage, metal–hydride hydrogen atom transfer (MHAT) has recently gained attention, as a promising strategy for radical hydrofunctionalization of unactivated alkenes2, thus enabling late-stage diversification of complex molecules3,4. However, owing to the weak interactions between the prochiral organic radical and the enantiopure catalyst5, asymmetric MHAT6 remains challenging. Here we show that cytochrome P450 enzymes (CYPs) can be repurposed to catalyse asymmetric MHAT, a new-to-nature reaction. Directed evolution of P450BM3 yielded a triple mutant that catalyses MHAT radical cyclization of unactivated alkenes, producing diverse cyclic compounds—including pyrrolidines and piperidines—with up to 98:2 enantiomeric ratio under aerobic whole-cell conditions. Apart from electron-deficient alkenes, alternative radical acceptors—including hydrazones, oximes and nitriles—were converted by repurposed P450BM3 to enantioenriched cyclization products. Mechanistic investigations support an MHAT mechanism proceeding by homolytic cleavage of a fleeting iron(III)–hydride species2,6. Starting from CYP119, directed evolution afforded a stereocomplementary MHATase, highlighting the potential of repurposed CYPs for MHAT biocatalysis. Our study highlights the prospect of integrating homolytic metal–hydride reactivity into metalloenzymes, thus expanding the scope of asymmetric radical biocatalysis.
