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科技导报  2015 

Salsolinol合成酶的克隆和表达

DOI: 10.3981/j.issn.1000-7857.2015.08.012, PP. 73-76

Keywords: Salsolinol,合成酶,定点突变,克隆表达,蛋白纯化

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Abstract:

Salsolinol合成酶是一种催化多巴胺和乙醛生成Salsolinol的酶,与帕金森病发病机制密切相关。研究发现Salsolinol合成酶与泛素的氨基酸序列高度相似,只有4个氨基酸位点有差异。本研究以泛素基因为模板,采用聚合酶链式反应技术对4个位点进行定点突变,将突变基因片段克隆到载体pET30a-GST上,构建pET30a-GST-Salsynthase重组载体,转化BL21后,IPTG诱导重组菌表达融合蛋白,经亲和层析柱纯化。结果表明,实现目的位点的定点突变,获得Sal合成酶基因,成功构建了GST-Salsynthase原核表达质粒,在大肠杆菌中表达纯化后得到较高纯度的GST-Salsynthase融合蛋白。

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