Abstract:

Laccase widely exists in soil and has potential industrial value in degradation of organic pesticide residues and lignin. In this study, a laccase gene, lac13H9, with total length of 1 389 bp was obtained from the soil macro genome. It encoded 462 amino acids polypeptide with a calculated molecular mass of 50 kDa and contained a signal peptide composed with 17 amino acid residues. The deduced amino acid sequence was a new protein through aligning with available protein sequences held in the NCBI. The recombinant lac13H9 heterologously expressed in Escherichia coli Transseta (DE3) and purified by the affinity chromatography on a Ni-NTA column. The maximal activity of laccase was observed at 40℃, the optimum pH was 6.0, and it had good thermal stability. About 60% enzyme activity was remained under 50℃ for 90 min. At the same time, low concentration of Fe2+ could promote laccase enzyme activity, while high concentration could inhibit enzyme activity. Fine enzymatic property of Lac13H9 has laid foundation for its application.

摘要：

漆酶广泛存在于土壤中，在有机农药残留和木质素的降解方面具有潜在的工业价值。从土壤宏基因组中得到了一个全长为1 389 bp漆酶基因lac13H9，编码462个氨基酸，预测理论分子量为50 kDa，含有一个17个氨基酸组成的信号肽，与NCBI蛋白数据库比对结果表明是一个新的漆酶基因。将lac13H9转入到大肠杆菌Transseta (DE3)中通过微好氧发酵进行异源表达，并通过Ni柱亲和层析纯化。获得的重组漆酶Lac13H9具有良好的酶学性质，其最适温度为40℃，最适pH为6.0，且具有较好的热稳定性，在50℃下保温90 min还有60%的剩余酶活力，同时发现低浓度的Fe2+促进漆酶酶活，高浓度则抑制酶活。良好的酶学性质为该酶的应用奠定了基础。

关键词: 土壤宏基因组；漆酶；基因克隆；酶学性质