Adsorption Characteristics of Affinity-ultrafiltration Escort to His-tagged Recombinant AxCeSD
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Abstract:
N-terminal histidine (His)6-tagged recombinant protein AxCeSD was purified by affinity chromatography. The affinity- ultrafiltration escort, Cu2+-IDA-Dextran T2000 specifically able to adsorb the His-tag of recombinant protein, was synthesized using water-soluble Dextran T2000 as the matrix (molecular weight 2000 ku), iminodiacetic acid as the chelating agent, and Cu2+ as the affinity ligand. The effects of pH, ionic strength, adsorption time, and reaction temperature on the adsorption characteristics of the escort with N-terminal His-tagged recombinant AxCeSD were investigated. The results showed that the adsorption capacity of the affinity-ultrafiltration escort for AxCeSD increased with the increases in pH and temperature within a certain range, but decreased as the ionic strength increased. Moreover, the adsorption equilibrium could be achieved within 30 min. The isotherm adsorption curve was fitted to the Langmuir model. The maximum adsorption yield was 125.15 mg/g and the dissociation constant was 3.259×10-6 mol/L, indicating that the specific adsorption of the affinity-ultrafiltration escort to recombinant AxCeSD was mainly from chelation.
