Clone, Expression and Characterization of the Thermostable β-Galactosidase from Soil Metagenomic DNA of Taishan Mountain

Abstract:

A possible β-Galactosidase gene (pwtsA) was discovered from soil metagenomic DNA of Taishan Mountain. PwtsA gene was inserted into the expression vector pET30a and transferred into E. coli BL21(DE3). Recombinant protein PWTSA was expressed as a soluble form at high level through IPTG induction, with a molecular mass of 57 kD analyzed by SDS-PAGE. PWTSA can produce o-nitrophenol from o-nitrophenol-β-D-galactopyranoside (ONPG), and its specific activity was determined as 13.6 U/mg. The enzymatic studies demonstrated that the recombinant protein PWTSA was a thermostable β-Galactosidase, its optimum temperature and pH were 85°C~95°C and 6.5 respectively. In standard assays, the Km for ONPG was 0.83 mmol/L.