Man-made Enzymes--from Design to in Vitro Compartmentalisation

In the past few years, a variety of methods have been developed to allow the in vitro evolution of a range of biomolecules including novel and improved biocatalysts (enzymes). These methods for directed evolution differ in the size and characteristics of the gene repertoire, in the way of linking genotype and phenotype, and in the selection approach. Selections for enzymes can be performed indirectly (for binding of a transition-state analogue or mechanism-based inhibitor), and directly using either intramolecular single-turnover selections (e.g. with SELEX) or the normal (intermolecular, multiple turnover) mode of enzymatic reactions. Each of these methods has distinct strengths and weaknesses. The best system (or combinations of systems) to use depends on the specific target for evolution and the evolutionary distance that needs to be crossed.