摘要
Enzymes are biocatalysts capable of catalyzing reactions under environmentally friendly conditions and can be applied in a wide variety of applications. However, biocatalysts are often not stable or not functional under the conditions required by industries. Therefore, natural enzymes must be engineered to overcome these problems. Apart from the directed evolution strategy, rational, semi-rational, and mechanistic-based enzyme engineering have made significant contributions to the field. Here, we discuss recently developed approaches that have been shown to be useful in enhancing enzyme capabilities. These examples have improved the thermostability, activity, and solubility of diverse enzymes by utilizing rational-design and semi-rational-design protein engineering in combination with computational tools and mechanistic data. We discuss weaknesses, strengths, and challenging points for each approach and highlight recent successful cases of enzyme engineering by these methods. We also discuss the current and emerging trends in enzyme engineering and the challenging points remaining to be addressed. Biocatalysis is a green and sustainable tool for production of fine chemicals. Enzyme engineering is important for fine-tuning enzymes to contain properties suitable for industrial biocatalysis. In addition to the well-known directed evolution strategy, rational-design and semi-rational-design protein engineering, which can be performed with just a small-sized library for screening, have emerged as promising methods for increasing enzyme stability, catalytic efficiency, selectivity, and substrate scope. We have observed growth in computational tools as well as the use of mechanistic information to guide enzyme engineering. This knowledge is valuable for improving enzyme performance with limited resources and small-sized libraries, which should allow a wider community to carry out successful enzyme engineering programs. Enzymes are versatile biocatalysts that can be used in a variety of applications. However, their inherent properties do not always meet industrial needs, making enzyme engineering necessary. Besides directed evolution, rational and semi-rational enzyme engineering have made important contributions toward improvement of enzyme efficiency. Here, we discuss recent approaches in improving enzyme thermostability, catalytic activity, and solubility through rational and semi-rational design in combination with computational tools and mechanistic data. Upcoming trends in these approaches are also discussed.