title: Octapeptide-based affinity chromatography of human immunoglobulin G: comparisons of three different ligands.
abstract: In an earlier work, we have developed a biomimetic design strategy based on the human IgG (hIgG)-Protein A interactions and identified an affinity ligand for hIgG, FYWHCLDE, which ranked top one in a pool of 14 potential candidates. Herein, two more octapeptides, FYCHWALE and FYCHTIDE, were identified, and the binding and purification of hIgG on the affinity columns packed with the three octapeptide-modified Sepharose gels were extensively studied and compared to find more effective octapeptide-based affinity ligands. It was found that all the three ligands bound hIgG and Fc fragment but barely bound Fab fragment, and the binding to hIgG and Fc was mainly by electrostatic interactions. The optimum binding pH values for the three ligands were different from each other, but kept in the range of 5.0-6.0. Ligand binding competition revealed that the binding sites on hIgG for the three octapeptides were similar to those for Protein A. Adsorption isotherms revealed that hIgG binding capacity was in the range of 64-104mg/mL drained gel in the order of FYWHCLDE>FYCHWALE>FYCHTIDE. Then, purifications of hIgG and human monoclonal antibody from human serum and cell culture supernatant, respectively, were achieved with the three affinity columns at high purities and recovery yields. Finally, the molecular basis for the binding affinity of the peptides for the Fc fragment of hIgG was elucidated by molecular dynamics simulations.
authors: Department of Biochemical Engineering and Key Laboratory of Systems Bioengineering of the Ministry of Education, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China.
authors: Wei-Wei
authors: WW
authors: Zhao
authors: Department of Biochemical Engineering and Key Laboratory of Systems Bioengineering of the Ministry of Education, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin 300072, China.
authors: Fu-Feng
authors: FF
authors: Liu
authors: Department of Biochemical Engineering and Key Laboratory of Systems Bioengineering of the Ministry of Education, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin 300072, China.
authors: Qing-Hong
authors: QH
authors: Shi
authors: Department of Biochemical Engineering and Key Laboratory of Systems Bioengineering of the Ministry of Education, School of Chemical Engineering and Technology, Tianjin University, Tianjin 300072, China; Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Tianjin 300072, China. Electronic address: ysun@tju.edu.cn.
authors: Yan
authors: Y
authors: Sun
mesh_terms: D000327:Adsorption; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001665:Binding Sites; D002846:Chromatography, Affinity / Q000379:methods*; D006801:Humans; D007074:Immunoglobulin G / Q000737:chemistry*; D008024:Ligands; D056004:Molecular Dynamics Simulation; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding; D013205:Staphylococcal Protein A / Q000378:metabolism; D055672:Static Electricity
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D008024:Ligands; D010455:Peptides; D013205:Staphylococcal Protein A
keywords: Affinity chromatography; Human immunoglobulin G; Molecular dynamics simulation; Monoclonal antibody; Peptide ligand; Purification