Strategies for discovery and improvement of enzyme function: state of the art and opportunities

Abstract

SummaryDevelopments in biocatalysis have been largely fuelled by consumer demands for new products, industrial attempts to improving existing process and minimizing waste, coupled with governmental measures to regulate consumer safety along with scientific advancements. One of the major hurdles to application of biocatalysis to chemical synthesis is unavailability of the desired enzyme to catalyse the reaction to allow for a viable process development. Even when the desired enzyme is available it often forces the process engineers to alter process parameters due to inadequacies of the enzyme, such as instability, inhibition, low yield or selectivity, etc. Developments in the field of enzyme or reaction engineering have allowed access to means to achieve the ends, such as directed evolution, de novo protein design, use of non‐conventional media, using new substrates for old enzymes, active‐site imprinting, altering temperature, etc. Utilization of enzyme discovery and improvement tools therefore provides a feasible means to overcome this problem. Judicious employment of these tools has resulted in significant advancements that have leveraged the research from laboratory to market thus impacting economic growth; however, there are further opportunities that have not yet been explored. The present review attempts to highlight some of these achievements and potential opportunities.

Putting bugs to the blush

“…14,15 Microorganisms have the advantage of high growth rates (and thus short production times) and readily scalable cultivation and production technologies. Furthermore, microbial production avoids the use of organic solvents, heavy metals, and strong acids or bases making processes more environmentally friendly as compared with chemical production.…”

“…Protein engineering can be a powerful approach in case individual enzyme parameters turn out to be limiting for the overall flux through the heterologous pathway. 15 Further optimization can be achieved through fine-tuning of individual promoters (constitutive or controlled induction of gene expression) and promoter strength. 16 Alternatively, all or selected genes of the new pathway can be expressed as operon under the control of a single promoter.…”

Biotechnological Applications of Proteases in Food Technology

“…Initially, they were mainly used as extracts of animal or vegetable tissues, or complete microorganism cells. The advances in enzyme isolation and production techniques have facilitated the direct application of purified enzymes (Schmid and others ; Kirk and others ; Kaul and Asano ; Li and others ). These technical advances permitted to extend the range of known enzymes and improve the characterization of their conformations and catalytic mechanisms, making their application simpler (Pogson and others ; Li and others ; Filici and others ; Strack ).…”

Directed evolution of an endoinulinase from Talaromyces purpureogenus toward efficient production of inulooligosaccharides

“…Therefore, there is a need for the development of efficient production processes of highly specific and active enzymes expressed in heterologous systems. This can be achieved by an adequate application of directed enzyme evolution . Herein, we describe the development of a recombinant expression system and application of directed enzyme evolution to engineer endoinulinases with increased expression yield in E. coli and increased specific activity compared to the wild type endoinulinase purified from the fungus Talaromyces purpureogenus ( Penicillium purpureogenum ATCC4713), annotated here as TPwt …”