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

A new method for synthesis of an Immobilized-Metal Affinity chromatography (IMAC) adsorbent with superparamagnetism (Fe₃O₄/SiO ₂-GPTMS-Asp-Co) was reported in this paper. Fe₃O ₄ nanoparticles were first modified by SiO₂ to form the core-shell Fe₃O₄/SiO₂ 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 Fe₃O₄/SiO₂-GPTMS-Asp nanoparticles with tetradentate ligands. Finally, the IMAC adsorbent with superparamagnetism, Fe₃O₄/SiO₂-GPTMS-Asp-Co, was finally obtained by the coordination of Co²⁺ 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. Fe₃O₄/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.