链球菌蛋白G免疫球蛋白结合域C3区串联体的表达、筛选与初步应用

Expression, Screening and Preliminary Application of the Concatenation C3 Region of the Streptococcal Protein G Immunoglobulin Binding Domain

筛选并制备高效结合抗体的重组蛋白G亲和层析介质。以链球菌蛋白G C3区为结构单元,拼接形成3、4、5、6个重复C3片段的串联体,分别克隆至PET21,并在E.coli BL21(DE3)中表达。经Ni^(2+)纯化的重组蛋白分别偶联至Purose 4 Fast Flow制备亲和层析介质,比较介质的吸附情况,发现重组四串体对h IgG的吸附量最高,其静态饱和吸附量为66.79 mg×mL^(-1),动态吸附载量为35 mg×mL^(-1)介质,优于同类型商品介质。利用该介质分离纯化小鼠腹水和人血清中的IgG,纯度在95%左右,回收率不低于80%。

In order to screen and prepare recombinant protein G affinity resins with high efficient antibody binding, the C3 region of Streptococcus protein G was used as the structural unit to prepare different tandem repeats（3, 4, 5 and 6） of C3 fragments, which were respectively cloned into PET21 and expressed in E. coli BL21（DE3）. The recombinant proteins purified by Ni^（2＋） affinity chromatography were coupled onto Purose 4 Fast Flow to prepare affinity chromatography resins. The 4-tandem repeats resin exhibits the highest adsorption capacity to h IgG with static adsorption capacity of 66.79 mg×mL^（-1） and dynamic adsorption capacity of 35 mg×mL^（-1）, which is better than same type commercial resins. Purity of 95% and recovery over 80% can be obtained for mouse ascites and human serum purification using this resin.