QuickTip #4: yield of His-tagged proteins

Immobilized Metal Ion Affinity Chromatography (IMAC) is used for purification of His-tagged proteins. Ni-NTA based resins are the most commonly used as the Ni2+ ion is considered to have the strongest affinity to histidine-tagged proteins but stringent requirements on the yield of target protein can make it well worth the extra effort of trying another metal ion to optimize the purification process...

Under some circumstances, other transition metal ions such as Co2+, Cu2+, Fe2+ and Zn2+ may prove to be more suitable. This is, of course, just one of the factors affecting binding of the His-tagged protein to the metal ion. There’s also the structure and characteristics of the protein and the pH and composition of the binding buffer etc.

When purifying His-tagged proteins expressed in yeast and mammalian cells screening of the optimized metal ion or ligand is suggested. Since these expression systems are more complex, the host cell impurities are more complex in comparison with His-tagged proteins expressed in bacterial host cells.

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