It is often desirable to design a method of purification directly into a recombinant protein expression strategy. This is most often accomplished by expressing a target protein with an appended “tag” that permits its purification by affinity chromatography. Tags can be a short peptide that specifically interacts with an immobilized antibody or metal ion, or larger binding domains that interact with specific immobilized ligands.
An important consideration in using a protein-tagging strategy is whether or not the presence of a tag will affect the protein’s downstream applications. For example, the presence of a larger binding domain may permit more simple purification, but it might also adversely affect the biochemical function of the target protein it is fused to. Therefore, it is often desirable to include a method for tag removal following purification of the fusion protein. Common methods include the use of site-specific proteases to cleave at an engineered site between the tag and the target protein, or the use of an intein to auto-catalyze tag removal.