Tanabe Y, Noguchi K, Morikawa A, Mizuno D, Soma G
Biotechnology Research Center, Teikyo University, Kawasaki 2, Japan.
Biochem Biophys Res Commun. 1991 Aug 30;179(1):683-8. doi: 10.1016/0006-291x(91)91426-d.
To study its biological functions, tumor necrosis factor precursor (proTNF) with a molecular size of 26-KDa was obtained as a recombinant protein from Escherichia coli. The recombinant proTNF was successfully accumulated in the insoluble form, corresponding to about 10-15% of total E. coli proteins. Solubilization, gel filtration and anion exchange chromatography were performed under denatured conditions followed by dialysis in phosphate-buffered saline. These processes removed most of the contaminating bacterial proteins, yielding proTNF with a purity of about 70-80%. This recombinant proTNF is expected to be useful for functional studies on activated macrophages with membrane integrated proTNF.
为研究其生物学功能，从大肠杆菌中获得了分子大小为26千道尔顿的肿瘤坏死因子前体（proTNF）作为重组蛋白。重组proTNF成功以不溶性形式积累，约占大肠杆菌总蛋白的10 - 15%。在变性条件下进行溶解、凝胶过滤和阴离子交换色谱，随后在磷酸盐缓冲盐水中透析。这些过程去除了大部分污染性细菌蛋白，得到纯度约为70 - 80%的proTNF。预计这种重组proTNF可用于对具有膜整合proTNF的活化巨噬细胞进行功能研究。
