Engineering P450 Peroxygenase Overoxidation Promiscuity Enables Concise and High-Yielding Biocatalytic Synthesis of d-Amino Acids from Fatty Acids

Direct asymmetric α-C−H bond amination of carboxylic acids is an attractive route to optically active α-amino acids from inexpensive, renewable starting materials. However, traditional chemical methods suffer from expensive catalysts and environmental concerns, while recently developed multienzyme cascades are limited by complex auxiliary enzyme requirements and narrow substrate scope. Here, we report an engineered P450 peroxygenase (P450CLA-M5) with enhanced overoxidation promiscuity that enables the direct conversion of fatty acids to α-keto acids. Coupling this enzyme with d-amino acid dehydrogenase and formate dehydrogenase establishes a biocatalytic cascade for the C−H amination of fatty acids to d-amino acids. Saturated, unsaturated, and aromatic fatty acids were converted to the corresponding d-amino acids with yields up to 94% and >99% ee. Specifically, the yield of d-2-aminodecanoic acid represents a 15-fold improvement over previous biocatalytic approaches. This work demonstrates how engineering increased enzyme promiscuity can enhance cascade efficiency for the synthesis of valuable chiral d-amino acids from renewable feedstocks.