One‐step selective affinity purification and immobilization of His‐tagged enzyme by recyclable magnetic nanoparticles

Abstract

Abstract The NiFe2O4 magnetic nanoparticles (NF‐MNPs) were prepared for one‐step selective affinity purification and immobilization of His‐tagged recombinant glucose dehydrogenase (GluDH). The prepared nanoparticles were characterized by a Fourier‐transform infrared spectrophotometer and microscopy. The immobilization and purification of His‐tagged GluDH on NF‐MNPs were investigated. The optimal immobilization conditions were obtained that mixed cell lysis and carriers in a ratio of 0.13 in pH 8.0 Tris‐HCl buffer at 30℃ and incubated for 2 h. The highest activity recovery and protein bindings were 71.39% and 38.50 μg mg–1 support, respectively. The immobilized GluDH exhibited high thermostability, pH‐stability and it can retain more than 65% of the initial enzyme after 10 cycles for the conversion of glucose to gluconolactone. Comparing with a commercial Ni‐NTA resin, the NF‐MNPs displayed a higher specific affinity with His‐tagged recombinant GluDH.

Keywords

• glucose dehydrogenase
• His‐tagged recombinant protein
• immobilization
• purification