Facile fabrication of nickel immobilized on magnetic nanoparticles as an efficient affinity adsorbent for purification of his-tagged protein

摘要

In the present research, an efficient, convenient and inexpensive method for the one-pot synthesis of Fe_3O_4@His-tidine is developed. Histidine is readily loaded on magnetic nanoparticles by one step and simple method without any supplemental linkers. In the structure of Fe_3O_4@Histidine, histidine covalently immobilized on the surface of Fe_3O_4, magnetic nanoparticles are able to trap Ni~(2+) ions through a strong interaction between nickel and histi-dines in protein tag. Two coordination sites of nickel are occupied with ligand on the surface of magnetic nanoparticles and four coordination sites have been remained that these sites will be occupied with histidine tag of recombinant protein A. The functionalized nanoparticles were spherical and well separated with an average diameter around 30 nm. The obtained magnetic nanoparticles have a saturation magnetization of about 54 emu/g. Fe_3O_4@Histidine-Ni was used to enrich and purify 6 × histidine-tagged recombinant protein-A directly from the mixture of lysed cells. It has been found that Ni(II)-immobilized Fe_3O_4@Histidine magnetic nanoparticles present negligible nonspecific protein adsorption and high His-tag protein binding capacity The average binding capacity (MW 42 k Da), is 700 ± 25 μg·mg~(-1) (protein/Fe_3O_4@Histidine-Ni).