Cloning and Expression of Sulfur Oxygenase Reductase Gene from Acidithiobacillus caldus and the Study of the Recombinant Enzyme Activity

doi:10.13560/j.cnki.biotech.bull.1985.2015.11.024

Abstract

Abstract: In order to investigate biochemical properties of sulfur oxygenase reductase from Acidithiobacillus caldus(AcSOR), the sor gene was amplified by PCR with the extracted A. caldus TST3 genomic DNA as template, which was inserted into vector pET-28a to construct recombinant plasmid pET-sor. And the plasmid was then transformed into Escherichia coli BL21(DE3)to obtain recombinant strain E.coli BL21(pET-sor2). SDS-PAGE showed that the target enzyme SOR could be expressed in this recombinant strain after induction by IPTG. Induction conditions were optimized. The recombinant AcSOR was purified with Ni⁺-NTA column from the supernatant of the sonicated E.coli BL21(pET-sor2)induced under the optimal conditions. For the oxidation reaction, specific activity, K_m and V_max of the purified AcSOR were 0.70 units of oxidase/mg of protein, 15.672×10^-2g/mL and 12.755×10^-5 mol/(L·min), respectively. Furthermore, specific activity, K_m and V_max of AcSOR for the reduction reaction were 2.21 units of reductase/mg of protein, 0.507×10^-2 g/mL and 4.876×10^-5 mol/(L·min), respectively.

Key words: bioleaching, Acidithiobacillus caldus, sulfur oxygenase reductase, enzyme activity

Li Lingling, Lü Zaosheng, Zuo Zhenyu, Li Yaoyi. Cloning and Expression of Sulfur Oxygenase Reductase Gene from Acidithiobacillus caldus and the Study of the Recombinant Enzyme Activity[J]. Biotechnology Bulletin, 2015, 31(11): 186-194.

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