Preparation and enzymatic behavior of surfactant-enveloped enzymes for glycosynthesis in nonaqueous aprotic media

Abstract

Abstract Surfactant-enveloped enzymes (SEEs) were prepared from pure cellulases, cellobiohydrolase I and endoglucanase I (Cel7A and Cel7B, respectively), via simply freeze-drying water-in-oil emulsions, wherein the aqueous phase containing each cellulase was stabilized with the nonionic surfactant, dioleyl- N - d -glucona- l -glutamate. The enzymatic tolerance of SEEs to various nonaqueous solvents was investigated, aiming at a novel synthetic approach in biocatalytic glycoengineering. SEE-Cel7A preserved ca . 67% of the original activity after 3 h incubation in lithium chloride (LiCl)/dimethylacetamide (DMAc) that is a good solvent for carbohydrates but completely deactivates intact enzymes. This excellent enzymatic durability depended on the preparation conditions of SEEs, e.g. pH and salt species of the aqueous phase during SEE preparation. SEE-Cel7A or SEE-Cel7B was applied as a biocatalyst to synthesize cellulose, a sugar polymer which is insoluble in common solvents but dissolves in LiCl/DMAc. Both SEEs could catalyze the direct dehydration of cellobiose without any activation of the anomeric carbon, a property that is indispensable for conventional chemo-enzymatic synthesis. The SEE-Cel7A provided short-chain cellulose with the degree of polymerization (DP) ca . 20, and longer-chain cellulose with DP ca . 60 was preferentially obtained by the SEE-Cel7B, possibly through preferential reverse hydrolysis instead of inherent hydrolysis. Nonaqueous SEE-mediated biocatalysis using inexpensive glycohydrolases and sugars that do not need to be chemically modified beforehand would have potentially wide applications in glycoengineering.