Directed evolution for enzyme development in biocatalysis.

Abstract As an important sector of the chemical industry, biocatalysis requires the continuous development of enzymes with tailor-made activity, selectivity, stability, or tolerance to unnatural environments. This is now routinely achieved by directed evolution based on iterative cycles of genetic diversification and activity screening. Here, we highlight its recent developments. First, the design of “smarter” libraries by focused mutagenesis may be a crucial start-up for a fast and successful outcome. Then library assembly and expression are also key steps that benefits from modern molecular biology progresses. Finally, various strategies may be considered for library screening depending on the final objective: while low-throughput direct assays have been very successful in generating enzymes for important biocatalytic processes, even in bringing completely new chemistries to the enzyme world, ultrahigh-throughput screening methods are emerging as powerful approaches for engineering the next generation of industrial enzymes.