Sugita Y, Nakano Y, Tomita M
Department of Physiological Chemistry, School of Pharmaceutical Sciences, Showa University, Tokyo.
J Biochem. 1988 Oct;104(4):633-7. doi: 10.1093/oxfordjournals.jbchem.a122524.
A protein which inhibited complement channel formation was isolated from extracts of papain-digested human erythrocyte membranes using DEAE-Sephacel, Bio-Gel A0.5m column chromatographies, and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by transfer to nitrocellulose paper and elution with 2% NP-40 solution. The purified protein showed a molecular weight of 18 kDa, and efficiently inhibited hemolysis of EC5-7 cells with C8 and C9, but did not show any decay-accelerating activity to C5 convertase. Immunochemical analysis of native membranes after sodium dodecyl sulfate-polyacrylamide gel electrophoresis using the antibody against this protein gave a single band having the same mobility as this protein; papain did not eliminate a significant portion of this protein.
使用DEAE-葡聚糖凝胶、Bio-Gel A0.5m柱色谱法,以及制备性十二烷基硫酸钠-聚丙烯酰胺凝胶电泳,随后转移至硝酸纤维素纸并用2% NP-40溶液洗脱,从木瓜蛋白酶消化的人红细胞膜提取物中分离出一种抑制补体通道形成的蛋白质。纯化后的蛋白质分子量为18 kDa,能有效抑制C8和C9对EC5-7细胞的溶血作用,但对C5转化酶没有任何促衰变活性。使用针对该蛋白质的抗体对十二烷基硫酸钠-聚丙烯酰胺凝胶电泳后的天然膜进行免疫化学分析,得到一条迁移率与该蛋白质相同的单一带;木瓜蛋白酶并未去除该蛋白质的大部分。