Assembling of engineered IgG-binding protein on gold surface for highly oriented antibody immobilization.
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.
Kanno
S
S
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
Yanagida
Y
Y
Haruyama
T
T
Kobatake
E
E
Aizawa
M
M
D000906:Antibodies / Q000378:metabolism*; D000941:Antigens / Q000378:metabolism; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D006046:Gold; D007074:Immunoglobulin G / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid; D013205:Staphylococcal Protein A / Q000235:genetics / Q000378:metabolism*
D000906:Antibodies; D000941:Antigens; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A; D006046:Gold; D003545:Cysteine