Directed evolution of carbon–hydrogen bond activating enzymes

[Display omitted] •C–H activating enzymes hold promise for a wide range of industrial applications.•New families of C–H activating enzymes offer desirable synthetic capabilities.•Low throughput screening still dominates C–H activating enzyme evolution campaigns.•Combinatorial use of advanced engineering technologies will accelerate evolution. As industrial biocatalysis is maturing, access to enzymatic activities beyond chiral resolutions, asymmetric ketone reductions and reductive aminations is gradually becoming reality. Especially the utilization of carbon–hydrogen bond (C–H) activating enzymes is very attractive as they catalyze a variety of chemically extremely challenging transformations. Because of their intrinsic complexity, the use of these enzymes in manufacturing has been limited. However, recent advances in enzyme engineering and bioinformatics have led to activity improvements for native and non-native substrates, the introduction of new-to-nature chemistries and the identification of promising novel enzyme families. Looking forward, the use of automation and advanced computer algorithms will help to streamline the evolution process of C–H activating enzymes leading to more robust and active biocatalysts.