Characterization of novel α-galactosidase in glycohydrolase family 97 from Bacteroides thetaiotaomicron and its immobilization for industrial application

Bacteroides thetaiotaomicron (B. thetaiotaomicron), which resides in the human intestinal tract, has a number of carbohydrate enzymes, including glycoside hydrolase (GH) family 97. Only a few GH 97 enzymes have been characterized Iodate. In this study, a novel alpha-galactosidase (Bt_3294) was cloned from B. theiaiotaomicron, expressed in Escherichia coli, and purified using affinity chromatography. This novel enzyme showed optimal activity at 60 degrees C and pH 7.0. Enzyme activity was reduced by 94.4% and 95.7% in the presence of 5 mM Ca2+ and Fe2+, respectively. It is interesting that Bt_3294 specifically hydrolyzed shorter a-galactosyl oligosaccharides, such as melibiose and raffinose. The D-values of Bt_3294 at 40 degrees C and 50 degrees C were about 107 and 6 mM, respectively. After immobilization of Bt_3294, the D-values at 40 degrees C and 50 degrees C were about 37.6 and 29.7 times higher than those of the free enzyme, respectively. As a practical application, the immobilized Bt_3294 was used to hydrolyze raffinose family oligosaccharides (RFOs) in soy milk, decreasing the RFOs by 98.9%. (C) 2020 Elsevier B.V. All rights reserved.