Immobilized-metal affinity chromatography adsorbent with paramagnetism and its application in purification of histidine-tagged proteins

Abstract

A new method for synthesis of an Immobilized-Metal Affinity chromatography (IMAC) adsorbent with superparamagnetism (Fe 3O 4/SiO 2-GPTMS-Asp-Co) was reported in this paper. Fe 3O 4 nanoparticles were first modified by SiO 2 to form the core–shell Fe 3O 4/SiO 2 with superparamagnetism, the core–shell microspheres were then successively treated by 3-glycidoxypropyltrimethoxysilane (GPTMS), l-aspartic acid ( l-Asp) and 2-bromoacetic acid to form Fe 3O 4/SiO 2-GPTMS-Asp nanoparticles with tetradentate ligands. Finally, the IMAC adsorbent with superparamagnetism, Fe 3O 4/SiO 2-GPTMS-Asp-Co, was finally obtained by the coordination of Co 2+ with the resulting nanoparticles. The intermediates and product obtained from the process mentioned above were characterized by TEM, SEM, XRD, XPS, FT-IR, TGA, AAS, EDS, element analysis and magnetic hysteresis loop. Fe 3O 4/SiO 2-GPTMS-Asp-Co nanoparticles used as IMAC adsorbent to separate the recombinant 6-histidine-tagged gp41 protein were investigated. The result indicates that the IMAC adsorbent we prepared has outstanding advantages in the separation of the 6-histidine-tagged proteins from the crude bacterial lysate, such as simple operation, high selectivity and capacity.