嗜热菌Bacillus sp. SCSIO 15121是从南中国海沉积物中分离的，本研究从其基因组中克隆了一个编码超氧化物歧化酶的基因SODBa3，该基因长度为609 bp，对应着202个氨基酸残基。本研究构建pET28a(+)-SODBa3表达载体，并在大肠杆菌BL21 (DE3)中实现了SODBa3的可溶性异源表达。本文研究了重组SODBa3的酶学性质，最适pH为8.0~8.5，最适反应温度为60 ℃，酶活为3215.6 U/mg。经鉴定，SODBa3对CH3Cl-C2H5OH敏感，但对H2O2不敏感，因此SODBa3是一种Mn离子特异的SOD。采用圆二色谱结合不同温度下酶活的变化研究酶的热稳定性，结果说明SODBa3在40~70 ℃处理1 h酶活保持较高的稳定性，在100 ℃下处理1 h剩余酶活为50%。金属离子耐受性试验表明5 mM的Mn2+对SODBa3的酶活有促进作用，SODBa3对乙醇和DMSO的耐受性较好，剩余酶活分别为对照的112.56±9.77和98.55±6.47%，这些结果表明SODBa3具有较好的工业应用前景。

An SOD gene (SODBa3) was cloned from the genome of Bacillus sp. SCSIO 15121 that was originally isolated from the sediment of the South China Sea. SODBa3 contains 609 base pairs (bp) and corresponds to 202 amino acid residues. The pET28a (+)-SODBa3 expression vector was constructed in this study, and heterologous expression of soluble SODBa3 was achieved in Escherichia coli BL21 (DE3). Enzymatic properties of the recombinant SODBa3 were then studied; the optimal pH and reaction temperature were 8.0~8.5 and 60 ℃, respectively, and the specific activity of SODBa3 was 3215.6 U/mg. The results showed that the activity of SODBa3 was sensitive to CH3Cl-C2H5OH but was not affected by hydrogen peroxide, indicating that SODBa3 is a Mn2+-dependent enzyme. The thermostability of SODBa3 was also studied by circular dichroism spectroscopy and by examination of changes in the activity of SODBa3 at various temperatures. The results revealed that SODBa3 was relatively stable during treatment at 40~70 ℃ for one hour, and the residual activity was 50% after incubation at 100 ℃ for one hour. The metal ion tolerance experiment indicated that 5 mM Mn2+ had stimulating effects on SODBa3 activity; SODBa3 showed good tolerance to 10% ethanol and dimethyl sulfoxide (DMSO), and the residual activities were 112.56%±9.77% and 98.55%±6.47% of the control activity, respectively. These findings indicate that microbial SODBa3 has high potential for utilization in the industry.

中国科学院战略性先导科技专项(XDA11030404)