Phosphorylated-tyrosine based pseudobioaffinity adsorbent for the purification of immunoglobulin G.
The present study evaluated the phosphorylated-tyrosine (P-Tyr) based pseudobioaffinity adsorbent for the purification of human immunoglobulin G (IgG). P-Tyr was selected as a ligand to mimic the natural interactions that occur between the immunoreceptor tyrosine-based activation motif and the IgG. The ligand was coupled to bisoxirane-activated agarose gel and the effect of buffer system, pH, and conductivity was performed to elucidate the nature of IgG-P-Tyr interactions. P-Tyr-agarose was able to purify IgG from human plasma solution in HEPES buffer at pH 7.0 exhibiting a purification factor of 9.1 with IgG purity of 91% (based on ELISA analysis of albumin, transferrin, and immunoglobulins A, G, and M). The evaluation of different functional groups of P-Tyr on the adsorption of human IgG indicated the predominance of electrostatic interactions with phosphate groups, although the contributions of aromatic and carboxylic groups also play a role. The thermodynamic parameters (ΔH°, ΔS°, ΔG°) for IgG adsorption onto P-Tyr-agarose were determined from the temperature dependence. The maximum IgG binding capacity at 20°C was 273.51±12.63mgg-1and the dissociation constant value of the complex IgG-P-Tyr was in the order of 10-5molL-1indicating low-affinity.
School of Chemical Engineering, University of Campinas (UNICAMP), 13083-970, Campinas, SP, Brazil.
Gisele Luiza
GL
Pavan
Chemical Engineering Department, Federal University of São Paulo (UNIFESP), 09913-030, Diadema, SP, Brazil.
Igor Tadeu
IT
Lazzarotto Bresolin
School of Chemical Engineering, University of Campinas (UNICAMP), 13083-970, Campinas, SP, Brazil.
Angélica
A
Grespan
School of Chemical Engineering, University of Campinas (UNICAMP), 13083-970, Campinas, SP, Brazil. Electronic address: sonia@feq.unicamp.br.
Sonia Maria
SM
Alves Bueno
D000327:Adsorption; D002021:Buffers; D002846:Chromatography, Affinity / Q000379:methods*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000302:isolation & purification*; D008024:Ligands; D019000:Phosphotyrosine / Q000737:chemistry*; D011485:Protein Binding; D012685:Sepharose / Q000737:chemistry; D013816:Thermodynamics
D002021:Buffers; D007074:Immunoglobulin G; D008024:Ligands; D019000:Phosphotyrosine; D012685:Sepharose
Adsorption; Human immunoglobulin G; Human plasma; O-Phospho-tyrosine; Purification