title: Immobilized protein ZZ, an affinity tool for immunoglobulin isolation and immunological experimentation.
abstract: 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 (DE3) 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 4B with butane-1,4-diol diglycidyl ether. The protein ZZ-Sepharose 4B 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.
authors.affiliation: State Key Laboratory of Pharmaceutical Biotechnology, College of Life Sciences, Nanjing University, Nanjing 210093, People's Republic of China.
authors.forename: Cheng
authors.initials: C
authors.lastname: Chen
authors.forename: Qi-Lai
authors.initials: QL
authors.lastname: Huang
authors.forename: Shu-Han
authors.initials: SH
authors.lastname: Jiang
authors.forename: Xiao
authors.initials: X
authors.lastname: Pan
authors.forename: Zi-Chun
authors.initials: ZC
authors.lastname: Hua
mesh_terms: D002072:Butylene Glycols / Q000737:chemistry; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D003432:Cross-Linking Reagents; D006639:Histidine / Q000235:genetics; D007074:Immunoglobulin G / Q000302:isolation & purification*; D047468:Immunoprecipitation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification; D012685:Sepharose / Q000737:chemistry; D013205:Staphylococcal Protein A / Q000737:chemistry*
chemical_list: D002072:Butylene Glycols; D003432:Cross-Linking Reagents; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A; D006639:Histidine; D012685:Sepharose; C014376:1,4-bis(2,3-epoxypropoxy)butane