**5. Conclusions**

We demonstrated that Amy A can perform both hydrolysis and alcoholysis in media containing deep eutectic solvents (DES) using starch or maltotriose as substrates. However, the enzyme activity and stability in each medium depends on the nature of the DES. The possibility of reducing water activity using alternative reaction media as the DES cosolvent was successful to a certain degree, as shown by the increase in the alcoholysis/hydrolysis ratio; however, it was found that using pure DES had a destabilizing effect. The alcholysis yield measured as methyl-glucoside with the optimal DES cosolvent was 20% greater than that previously reported in an aqueous buffer with the same methanol concentration.

Our results highlight the importance of performing variable temperature structural studies to determine the temperature limit at which an enzyme is operative.

**Supplementary Materials:** Supplementary materials can be found at http://www.mdpi.com/1422-0067/20/21/ 5439/s1.

**Author Contributions:** Conceptualization, G.S.-R., E.C. and A.L.-M.; methodology, A.M.-M. W.X. R.A.-B. S.S., L.O.; validation, W.X.; writing—original draft preparation, A.M.-M. W.X. R.A.-B. G.S.-R. funding acquisition, G.S.-R.

**Funding:** This research was funded by Dirección General de Asuntos del Personal Académico, Universidad Nacional Autónoma de México through PAPIIT Grants No. IT200617 to G.S.R. and IA202619 to W.X.

**Acknowledgments:** The authors thank Humberto Flores and Fernando González-Muñoz for technical assistance. Juan Manuel Hurtado and the following facilities of the Instituto de Biotecnología, Universidad Nacional Autónoma de México for their services at different stages of this work: Unidad de Cómputo, Unidad de Síntesis y Secuenciación and Unidad de Biblioteca.

**Conflicts of Interest:** The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

#### **References**

1. Larsson, J.; Svensson, D.; Adlercreutz, P. α-Amylase-catalysed synthesis of alkyl glycosides. *J. Mol. Catal. B Enzym.* **2005**, *37*, 84–87. [CrossRef]


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