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

The His-tag plays a crucial role in the purification process of proteins like Green Fluorescent Protein (GFP). A His-tag is a sequence of histidine residues that is often added to the N- or C-terminus of a protein, which in this case is attached to GFP. This tag facilitates the purification of the protein through a process known as affinity chromatography.

In the purification process, the His-tag specifically binds to nickel or cobalt ions that are immobilized on a chromatography column. When the cell lysate containing the His-tagged protein is passed through this column, the protein of interest (in this scenario, GFP) binds to the metal ions while other non-target proteins wash away. Subsequently, the bound GFP can be eluted from the column by using a solution containing imidazole, which competes with the His-tag for binding to the metal ions.

This selective binding and elution process significantly simplifies the isolation of GFP, allowing researchers to obtain a high-purity sample of the protein efficiently. Thus, the role of the His-tag in GFP purification is centered on its ability to enable selective purification, making it a fundamental tool in protein biochemistry.