摘要
在大肠杆菌中表达的重组Hepcidin融合蛋白以包涵体形式存在,其N端带有6个组氨酸。以Ni2+-IDA-Sepharose Fast Flow为层析介质,在变性条件下以不同的咪唑和pH值洗脱方式对Hepcidin融合蛋白的纯化效果进行了比较,确定了该融合蛋白的金属螯合层析纯化条件。以60 mmol/L咪唑洗脱杂蛋白,然后将pH值降至4.0洗脱融合蛋白,纯化后的融合蛋白纯度大于95%,而且不含咪唑,有利于下一步Hepcidin的制备。金属螯合层析中融合蛋白收率不低于90%。Ni2+-IDA-Sepharose Fast Flow对该融合蛋白的吸附量为30.4 mg/mL。
Hepcidin fusion protein having a N-terminal hexahistidine tag was expressed in E. coli as inclusion bodies. In this paper, different pH and imidazole elution methods were investigated for the purification of Hepcidin fusion protein by immobilized metal ion affinity chromatography under denaturation conditions. At last, two-step elution was used for purification of fusion protein. After contaminant proteins were removed by 60 mmol/L imidazole, the fusion protein fraction having a purity of 〉95 % was eluted with pH4.0 buffer. Moreover, the purified protein didn＇ t contain imidazole and had a high concentration of protein. The binding capacity of Ni^2 ＋ -IDA Sepharose Fast Flow for Hepcidin fusion protein was about 30.4 mg/mL resin, and the recovery of Hepcidin fusion protein was no less than 90 %.
出处
《北京化工大学学报（自然科学版）》
CAS
CSCD
北大核心
2005年第6期15-19,共5页
Journal of Beijing University of Chemical Technology(Natural Science Edition)
