Kanno S, Yanagida Y, Haruyama T, Kobatake E, Aizawa M
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
J Biotechnol. 2000 Jan 21;76(2-3):207-14. doi: 10.1016/s0168-1656(99)00186-8.
The B-domain, which is one of IgG-binding domains of staphylococcal protein A, was repeated five times and a cysteine residue was introduced at its C-terminus by a genetic engineering technique. The resulting protein, designated B5C1, retained the same IgG-binding activity as native protein A. The B5C1 was assembled on a gold plate surface by utilizing a strong affinity between thiol of cysteine and a gold surface. IgG-binding activity of B5C1 on a gold surface was much higher than that of physically adsorbed B5, which lacks cysteine residue. Furthermore, antigen-binding activity of immobilized antibody molecules through the use of assembled B5C1 on a gold surface was about 4.3 times higher than that of physically adsorbed antibody molecules. Immobilization of highly oriented antibody molecules was realized with the engineered IgG-binding protein.
B结构域是葡萄球菌蛋白A的IgG结合结构域之一，重复了5次，并通过基因工程技术在其C末端引入了一个半胱氨酸残基。所得蛋白质命名为B5C1，保留了与天然蛋白A相同的IgG结合活性。利用半胱氨酸的硫醇与金表面之间的强亲和力，将B5C1组装在金板表面。B5C1在金表面的IgG结合活性远高于缺乏半胱氨酸残基的物理吸附的B5。此外，通过在金表面组装B5C1固定化的抗体分子的抗原结合活性比物理吸附的抗体分子高约4.3倍。利用工程化的IgG结合蛋白实现了高度定向抗体分子的固定化。
