Heterologous Expression and Enzymatic Properties of Recombinant β-Galactosidase from Microbulbifer sp. ALW1
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Abstract:
β-Galactosidase, which catalyzes the hydrolysis of lactose into glucose and galactose, is one of the most important enzymes used in dairy processing. In this study, the β-galactosidase from Microbulbifer sp. ALW1 was heterologously expressed and purified in Escherichia coli BL21 (DE3), and the enzymatic properties of the purified enzyme were examined. The results showed that the β-galactosidase from Microbulbifer sp. ALW1 belonged to the glycoside hydrolase (GH) family 1. The recombinant β-galactosidase obtained by Ni-NTA agarose affinity chromatography had a molecular weight of 64 ku. The optimal reaction temperature and pH of the recombinant β-galactosidase were 30 ℃ and 4.5, respectively. The β-galactosidase exhibited good stability when the temperature was lower than 25 ℃ and pH was in the range of 4.0~5.0. The recombinant β-galactosidase had good tolerance to DTT, Tween 20, and Tween 80. In the presence of ionic detergents, SDS and CTAB, β-galactosidase almost lost its activity. The catalytic constants Km and Vmax of the recombinant β-galactosidase were 10.98 mmol/L and 7.48 U/mg, respectively. Structure simulation studies revealed that the catalytic acid/base residue was Glu186 and the nucleophilic residue was Glu370 for the β-galactosidase from Microbulbifer sp. ALW1. The results layed a theoretical foundation for the applications of β-galactosidase from Microbulbifer sp. ALW1 in the food field.
