重组人GM-CSF/MCAF融合蛋白的变性、复性及纯化研究

Denaturation,Renaturation and Purification of A Recombinant Human GM-CSF/MCAF Fusion Protein

人粒细胞巨噬细胞集落刺激因子(GM-CSF)和单核细胞趋化激活因子(MCAF)融合蛋白在大肠杆菌中高效表达后,表达产物以包涵体形式存在。包涵体经分离和洗涤后,探索了rhGM-CSF/MCAF变性和复性的合适条件。复性后的样品经Sephadex G-75凝胶过滤和CM-Sepharose FF离子交换两步层析,得到了具有生物学活性的SDS-PAGE纯的rhGM-CSF/MCAF。Western blot检测表明,纯化的rhGM-CSF/MCAF能分别与GM-CSF和MCAF抗体发生特异反应。

A fusion Protein of human graulocyte-macrophage stimulating factor (GM-CSF) and monocyte chemoattractant activating factor (MCAF) was efficiently expressed in Escherichia coli as inclusion bodies. After the isolatin of the inclusion bodies, the optimum conditions of denaturation and renaturation were studied. The resulting renatured rh GM-CSF/MCAF was purified to homogeneity by Sephadex G-75 gel-filtration and CM-Sepharose FF ion-exchange chromatography. The purified product remains biological activities and can react specifically with GM-CSF and MCAF antibodies respectively by western blot analysis.