An Analysis upon Various Strategies for Redesign and Direct Evolution of Enzyme Engineering | Original Article

ABSTRACT:

Enzymes are biocatalysts evolved in nature to achieve the speed and coordination of nearly all the chemical reactions that define cellular metabolism necessary to develop and maintain life. The application of biocatalysis is growing rapidly, since enzymes offer potential for many exciting applications in industry. The advent of whole genome sequencing projects enabled new approaches for biocatalyst development, based on specialised methods for enzyme heterologous expression and engineering. The engineering of enzymes with altered activity, specificity and stability, using sitedirected mutagenesis and directed evolution techniques are now well established. The use of enzymes in industrial applications has been recognised for providing clean processes with minimal impact on the environment. Enzyme engineering is undergoing the most profound and exciting transformation in its history. It promises unprecedented expansion in the scope and applications of modified or improved enzymes with desired physical and catalytic properties. Two complementary strategies are currently available: rational redesign and directed evolution. Although both approaches have met with great success, each has limitations. In this article, the perspectives for these enzyme-engineering strategies are discussed briefly. Directed evolution has become the preferred engineering approach to generate tailor-made enzymes. The method follows the design guidelines of nature: Darwinian selection of genetic variants. This review discusses the different stages of directed evolution experiments with the focus on developments in screening and selection procedures. Enzymes are increasingly being used in an industrial setting as a cheap and environmentallyfriendly alternative to chemical catalysts. In order to produce the ideal biocatalyst, natural enzymes often require optimization to increase their catalytic efficiencies and specificities under a particular range of reaction conditions. A number of enzyme engineering strategies are currently employed to modify biocatalysts, improving their suitability for large-scale industrial applications. These include various directed evolution techniques, semi-rational design techniques, and more recently, the de novo design of novel enzymes.