**5. Summary**

grinding aid, and milling frequency. The reaction features environmentally friendly, simple, and efficient operation. A major feature distinguishing this enzyme promiscuity from previously reported work is the use of mechanochemical ball milling techniques that overcome disadvantages such as long reaction times and the use of hazardous organic solvents. This work demonstrates the potential application of mixed enzyme-catalyzed reactions for drug synthesis under ball milling

In 2013, Berlozecki et al. reported for the first time an enzyme-catalyzed Ugi reaction that has many advantages over previous reactions, such as good reaction at room temperature and extensive solvent selection (**Figure 57**) [72]. In this threecomponent reaction, the aldehyde, amine, and isocyanide are condensed to form a dipeptide. Of all the selected lipases, Novozym 435 had the highest yield of 75%. Recently, Thomas et al. reported the concatenation of the Ugi four-component synthesis, and the CALB-catalyzed aminolysis of the intermediary formed Ugi methyl ester products furnishes a novel consecutive five-component reaction for the formation of triamides (**Figure 58**) [73]. This one-pot method is compatible with metal catalysis methods such as copper-catalyzed alkyl azide ring addition and

The mild reaction conditions make this sequence superior to the stepwise process with isolation of the Ugi product and even more favorable than other basecatalyzed or microwave-assisted aminolyses. This efficient scaffold forming

*Consecutive six-component U-4CR-CALB-catalyzed aminolysis-Suzuki cross-coupling sequence of biaryls.*

Suzuki cross-coupling or both in a one-pot process.

conditions.

**Figure 57.**

**Figure 58.**

**62**

*CALB-catalyzed Ugi reaction.*

**4.4 Ugi reaction**

*Molecular Biotechnology*

This chapter has reviewed some examples of various types of hydrolase catalytic promiscuous reactions and their applications in the past decade. Several different types of hydrolases catalyzed carbon-carbon or carbon-heteroatom formation reactions have been discussed: aldol reactions, Michael reactions, and multicomponent reactions.

From these examples, it is clear that enzymes that display catalytic promiscuity can provide new opportunities for organic synthesis. Exploiting enzyme catalytic promiscuous reactions might lead to new, efficient, and stable catalysts with alternative activity and could provide more promising and green synthetic methods for organic chemistry. The development of protein engineering and enzyme engineering can extend the application of metagenome libraries and find enzymes with specific promiscuous behavior. We believe the progress in the area of biocatalytic promiscuity will greatly extend the useful applications of enzymes.
