Principle

Protino® Ni-NTA products enable fast and convenient purification of recombinant polyhistidine-tagged proteins by immobilized metal ion affinity chromatography (IMAC). Proteins from any expression system can be purified under native or denaturing conditions. Binding of protein is based on the interaction between the polyhistidine tag of the recombinant protein and immobilized Ni²⁺ ions. Protino® Ni-NTA Agarose consists of the chelating ligand nitrilotriacetic acid (NTA) immobilized on 6 % cross-linked agarose beads that are suitable for batch binding, gravity-flow, and FPLC™ columns. The resin is precharged with Ni2+ ions and therefore ready to use. Protino® Ni-NTA Agarose uses NTA, which represents the most commonly used chelating ligand in IMAC. NTA is a tetradentate chelator which occupies four of the six binding sites in the coordination sphere of the Ni²⁺ ion. The remaining two coordination sites are usually occupied by water molecules and can be exchanged with histidine residues of the recombinant protein. This formation of coordination sites has turned out to be optimal for purification of polyhistidine-tagged proteins: Two available binding sites in the coordination sphere of the Ni²⁺ ion enable tight but reversible selective protein interactions. Chelation of Ni²⁺ ions by NTA through four coordination positions minimizes metal leaching during purification and increases specificity for polyhistidine-tagged proteins.

Features

* Binding capacity will vary for each polyhistidine-tagged protein
** High flow rates may reduce binding capacity