What is the role of a His-tag in the purification of GFP?

The His-tag plays a crucial role in the purification of GFP by facilitating affinity purification through its ability to bind to metal ions, typically nickel or cobalt immobilized on a resin. When a protein with a His-tag is passed through a column containing these metal ions, the His-tag interacts specifically with them, allowing for the selective capture of the tagged protein from a mixture of proteins. This targeting makes it much easier to isolate GFP from other cellular components and impurities, thus enhancing the purity and yield of the protein. After binding, the unbound impurities can be washed away, and the His-tagged GFP can be eluted by using a solution with a high concentration of imidazole, which competes with the His-tag for binding sites on the metal ions. This strategy is a well-established method in protein purification due to its efficiency and specificity.