β-glucosidase from Streptomyces griseus: Ester hydrolysis and alkyl glucoside synthesis in the presence of Deep Eutectic Solvents

Abstract

Abstract Deep Eutectic Solvents (DES) are ionic liquid analogs that have attracted considerable attention as green solvents for biocatalytic transformations. The use of DES as part of a ‘solvent engineering’ approach to enhance enzyme stability holds great promise since they are biodegradable, relatively inexpensive and environmentally safe media for enzyme reactions. However, the behaviour of specific enzymes in such solvents is complex; some enzymes are inhibited in DES, while others appear to be activated. Glucosidases are among the most widely used enzymes for commercial chemoenzymatic synthesis. In particular, their application in the synthesis of biodegradable alkyl glucosides by reverse hydrolysis is of great interest. Previous work in this laboratory identified Streptomyces griseus glucosidase (Sgβgl) as an interesting enzyme for biotechnological applications. In this study, we examined its behaviour in the presence of DES as a co-solvent using choline chloride as hydrogen bond donor and using glycerol, glucose and urea as hydrogen bond acceptors. We show that Sgβgl activity depends on both the nature of DES components, and their ratio in the eutectic mixture, as well as the water content of the reaction medium. A choline chloride/glycerol DES mixture at a level of 40% (v/v) caused activation of Sgβgl and increased its optimum temperature from 70 to 80\xa0\u200b°C: it also led to a striking increase in its thermostability, doubling its half-life at 60\xa0\u200b°C and almost tripling its half-life at 80\xa0\u200b°C. The synthesis of alkyl glucosides was explored using DES as a co-solvent. In the presence of DES, Sgβgl catalysed the formation of a range of alkyl glucosides. The presence of DES resulted in enhanced product yield, which was observed to increase with increasing temperature, up to 60\xa0\u200b°C. These studies show that the application of DES at relatively low % (v/v) levels can dramatically effect enzyme activity and stability. Specifically, enhanced thermostability can significantly increase the operating range for glucosidases for biocatalytic applications. Solvent engineering offers a simple and effective way to enhance glucosidase stability and will be useful as an alternative and/or adjunct to more complex methods such as immobilisation or protein engineering.