Abstract:

In order to find the ideal industrialized production of trehalose synthase gene engineering bacterium, the Tres gene was amplified by PCR with Changbai Mountain hot spring SH-110strain’s genomic DNA. The amplified Tres gene was cloned into prokaryotic expression vector pET-30a+ and expressed in BL21(DE3) under induction of IPTG and lactose. The expressed product was identified with SDS-PAGE and studied on characterization of the enzyme. The Tres gene at a length of 2912 bp was successfully cloned, in the experiments that joined IPTG and lactose, fusion protein in intracellular had been expressed, and added lactose protein expression induced by high volume. The relative molecular mass of expressed recombinant Tres was about 110 kDa. This enzyme exhibited the highest activity at the conditions of pH 7.0 and 60℃. Furthermore, metal ions such as Ca2+, K+, Zn2+ could activate the enzyme activity, while metal ions such as Ba2+, Cu2+, Mn2+decreased the enzyme activity. The kinetic parameters Km and Vmax were 8.823 mmol/L and 235.29 mmol/L. Recombinant Tres was expressed in E.coli, which laid the further study of recombinant gene-engineered strain produce large amounts of trehalose synthase provides theoretical basis for mass production.