6xHis-Ni-NTA Chromatography as a Superior Technique in Recombinant Protein Expressiod/Purification

Abstract

The 6xHis/Ni-NTA system is a fast and versatile tool for the affinity purification of recombinant proteins and antigenic peptides. It is based on the high-affinity binding of six consecutive histidine residues (the 6xHis tag) to immobilized nickel ions, giving a highly selective interaction that allows purification of tagged proteins or protein complexes from <1% to >95% homogeneity in just one step (1, 2). The tight association between the tag and the resin allows contaminants to be easily washed away under stringent conditions, yet the bound proteins can be gently eluted by competition with imidazole, or a slight reduction in pH. Moreover, because the interaction is independent of the tertiary structure of the tag, 6xHis labeled proteins can be purified even under the strongly denaturing conditions required to solubilize inclusion bodies.