Outstanding Performance of Novel Magnetic Beads for Efficient Purification of His-Tagged Proteins

Abstract

Placing of affinity tags at desired position of the proteins has been widely used to facilitate protein separation. One of the most commonly method for protein separation is nickel ion (Ni2+) affinity chromatography. Complex steps and time consuming in nickel ion affinity chromatography have caused high demand for alternative purification methods that allow direct and rapid separation of His-tagged proteins. In the present study, magnetic core–shell beads enriched with Ni2+-nitrilotriacetate (Ni-NTA) on their surface were prepared in this study by precipitation polymerization, with a uniform size of about 10 μm. The presented Ni2+ amount was 2.23%, which endowed the spheres with excellent magnetic responsivity and dispersity. We compared the magnetic beads enrichment method and nickel ion affinity chromatography method for extraction of His-tagged protein sample with different concentrations. The results revealed that, when compared with nickel ion affinity chromatography, extraction efficiency, purity of the extracted protein, and regeneration effort were much higher with the magnetic beads enrichment method. By using magnetic beads enrichment method, samples could be separated within 20 s, and purified within 1 h, with unlimited flow velocity and sample capacity. In addition, the magnetic beads enrichment method was particularly promising for the separation of His-tagged proteins in less samples.