Immobilized protein ZZ, an affinity tool for immunoglobulin isolation and immunological experimentation.
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.
Chen
Cheng
C
State Key Laboratory of Pharmaceutical Biotechnology, College of Life Sciences, Nanjing University, Nanjing 210093, People's Republic of China.
Huang
Qi-Lai
QL
Jiang
Shu-Han
SH
Pan
Xiao
X
Hua
Zi-Chun
ZC
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*
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