What is the role of imidazole in the purification of His-tagged GFP?

The role of imidazole in the purification of His-tagged GFP primarily involves its function as a competitive ligand. During the purification process, His-tagged proteins are typically bound to a metal affinity chromatography column, where the His-tag binds to metal ions, such as nickel or cobalt, that are immobilized on the resin. Imidazole, which has a similar structure to histidine, can compete with the histidine residues of the His-tag for binding sites on these metal ions.

When imidazole is introduced into the elution buffer, it effectively displaces the His-tagged protein from the metal ions on the column. By gradually increasing the concentration of imidazole, the purification process allows for the elution of the His-tagged protein in a controlled manner while reducing the binding of other non-specific proteins that may not have a His-tag. This selective elution significantly enhances the purity of the His-tagged protein, such as GFP, making imidazole a crucial component in the purification strategy.

The other options provided do not accurately reflect the specific role imidazole plays in the context of His-tagged protein purification. While stabilizing protein structure, enhancing fluorescence, or promoting bacterial growth are important aspects of working with proteins, they