Selective binding, magnetic separation and purification of histidine-tagged protein using biopolymer magnetic core-shell nanoparticles

Abstract In previous studies, we synthesized the magnetic core-shell structured Fe 3 O 4 /PMG/IDA-Ni 2+ nanoparticles. The Ni 2+ on the surface of nanoparticles provides abundant docking sites for histidine, and the composite nanoparticles showed potential applications in the separation and purification of histidine-tagged (His-tagged) proteins. Meanwhile, the presence of the superparamagnetic core (Fe 3 O 4 ) in the nanoparticles allows them to be quickly separated and purified by an external magnetic field. Herein, the ability of magnetic nanoparticles to purify His-tagged human superoxide dismutase 1 (hSOD1) was verified. SDS-PAGE and activity data showed His-tagged hSOD1 specifically bound to Fe 3 O 4 /PMG/IDA-Ni 2+ , and there was no significant competition for binding between final and three intermediate products. The binding capacity of nanoparticles can reach to 62.0 mg/g (dry weight of hSOD1/nanoparticles). The nanoparticle-bound hSOD1 exhibited better thermal and storage stability compared to free hSOD1. Furthermore, the purification efficiency of the magnetic nanoparticles in the separation and purification of His-tagged proteins was comparable to the other two commercial materials (High Affinity Ni-NTA Resin, HisPur Ni-NTA Magnetic Beads). Finally, the magnetic nanoparticles can be reused in the binding of His-tagged protein for multiple times. In conclusion, the nanoparticles are ready to be applied in the separation and purification of His-tagged protein.