|زهرا رشید,رامین قهرمان زاده,محمدرضا نژادمقدم,محبوبه نظری,محمدرضا شکری,حسین نعیمی,امیرحسن زرنانی
|J CHROMATOGR A
|ضریب تاثیر (IF)
|علمی - پژوهشی
|کشور محل چاپ
چکیده مقالهIn this research, a simple, efficient, inexpensive, rapid and high yield method for the purification of 6 × histidine-tagged recombinant protein was developed. For this purpose, manganese ferrite magnetic nanoparticles (MNPs) were synthesized through a co-precipitation method and then they were conveniently surface-modified with tetraethyl orthosilicate (TEOS) in order to prevent oxidation and form high density of hydroxyl groups. Next, the salen ligand was prepared from condensation reaction of salicylaldehyde and 3-aminopropyl (trimethoxy) silane (APTMS) in 1:1 molar ratio; followed by complexation with Ni(OAc)2.4H2O. Finally, the prepared Ni(II)-salen complex conjugated to silica coated MNPs and MnFe2O4@SiO2@Ni-Salen complex nanoparticles were obtained. The functionalized nanoparticles were spherical with an average diameter around 70 nm. The obtained MNPs had a saturation magnetization about 54 emu/g and had super paramagnetic character. These MNPs were used efficiently to enrich recombinant histidine-tagged (His-tagged) protein-A from bacterial cell lysate. In about 45 min, highly pure His-tagged recombinant protein was obtained, as judged by SDS-PAGE analysis and silver staining. The amount of target protein in flow through and washing fractions was minimal denoting the high efficiency of purification process. The average capacity of the matrix was found to be high and about 180 ± 15 mg g−1 (protein/MnFe2O4@SiO2@Ni-Salen complex). Collectively, purification process with MnFe2O4@SiO2@Ni-Salen complex nanoparticles is rapid, efficient, selective and whole purification can be carried out in only a single tube without the need for expensive systems.
tags: Magnetic nanoparticles Protein purification Immobilized metal affinity chromatography (IMAC) Histidine-tagged protein Salen-type ligands