Engineering peroxygenases for pharmaceutical applications: recent advances and future perspectives.

Selective oxidation is a cornerstone process in pharmaceutical biosynthesis, as it enables the conversion of inert C-H bonds into valuable functional groups such as alcohols, ketones, epoxides, and carboxylic acids, which serve as important intermediates in drug synthesis. Peroxygenases, including unspecific peroxygenases and P450 peroxygenases, have emerged as powerful biocatalysts for enzymatic oxidation, with growing applications in metabolite synthesis, late-stage C-H functionalization, and the preparation of enantioenriched intermediates for pharmaceutical development. In this review, we present the latest advances in peroxygenases engineering for pharmaceutical applications. We summarize pharmaceutical drug candidates that can serve as substrates for peroxygenases, discuss the enzymatic oxidation pathways mediated by peroxygenases for these substrates, and outline protein engineering strategies employed to improve the catalytic performance and properties of peroxygenases in pharmaceutical biosynthesis. We also discuss the potential of emerging artificial intelligence (AI) strategies for multi-dimensional peroxygenase engineering including catalytic activity, thermostability, hydrogen peroxide tolerance, heterologous expression, substrate scope, regioselectivity, and enantioselectivity. Overall, continued progress in peroxygenase engineering will drive the transformation of these biocatalysts for versatile industrial applications, thereby enabling sustainable drug synthesis, efficient metabolite production, and innovative therapeutic strategies.