Immobilized protein ZZ, an affinity tool for immunoglobulin isolation and immunological experimentation
Immobilized protein ZZ, an affinity tool for immunoglobulin isolation and immunological experimentation
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DOI：
10.1042/ba20060055
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发表时间：
2006-09-01
影响因子：
2.8
通讯作者：
Hua, Zi-Chun
中科院分区：
文献类型：
--
作者：
Chen, Cheng;Huang, Qi-Lai;Hua, Zi-Chun
目前，亲和纯化IgG的常用工具是固定化蛋白A，其从金黄色葡萄球菌的天然细胞壁组分中分离。天然蛋白A的制备复杂且昂贵。ZZ蛋白是一个人工合成的Fc区结合结构域，来源于A蛋白的B结构域。本研究在T7启动子的调控下，在BL 21（M）中表达了N端带有六聚组氨酸标签的重组ZZ蛋白。通过一步Ni 2+螯合亲和层析以50 mg蛋白质/升培养物的产量纯化蛋白质。然后用丁二醇二缩水甘油醚与Sepharose 413共价偶联。蛋白质ZZ-Sepharose 413树脂在IgG的亲和纯化中表现出良好的性能，以及在免疫沉淀实验中捕获蛋白质相互作用复合物。与天然蛋白A相比，ZZ蛋白的高效表达和纯化方法简单，成本低。因此，ZZ蛋白在免疫分析中的广泛应用是可行的，值得期待。
At present, the common tool for affinity purification of IgG is immobilized Protein A, which is separated from native cell-wall components of Staphylococcus aureus. It is complicated and costly to prepare natural Protein A. ZZ protein is a synthetic Fc region-binding domain originated from B domain of Protein A. In the present study, recombinant ZZ protein with a hexahistidine tag at the N-terminus was expressed in BL21 (M) under the control of T7 promoter. The protein was purified through one-step Ni2+ chelating affinity chromatography at a yield of 50 mg of protein/litre of culture. Then it was covalently coupled with Sepharose 413 with butane-1,4-diol diglycidyl ether. The protein ZZ-Sepharose 413 resin exhibited good performance in affinity purification of IgG, as well as in capturing the protein-interacting complexes in immunoprecipitation experiments. Compared with natural Protein A, the expression and purification of ZZ protein at high yield are very simple and low-cost. At this point, extensive applications of protein ZZ in immunoassays are practicable and to be anticipated.