Protein Cleavage Enzymes

In the process of recombinant protein expression, fusion proteins or polypeptide tags are usually constructed to facilitate the expression, tracing, and purification of recombinant proteins. Tags can be mainly divided into two types. One type is tags for affinity purification, such as 6His, GST, Strep-Tag, MBP, FLAG, and combined dual-tags or multi-tags. The other type is tags that increase protein solubility and assist in correct protein folding, such as GST, SUMO, MBP, TrxA, NusA, etc.

Among them, protein tags like GST, MBP, and SUMO have relatively large molecular weights and usually need to be removed by enzymatic cleavage to eliminate their influence on the structure and function of the target protein. Polypeptide tags ( such as 6*His, Strep-Tag, FLAG, etc. ) have small molecular weights and low immunogenicity, and basically do not affect the structure and function of proteins, so they usually do not need to be removed by enzymatic cleavage. However, for proteins used in structural biology or other advanced research, polypeptide tags with a certain molecular weight usually need to be removed by enzymatic cleavage to achieve the best results.

The method of removing tags lies in inserting the amino acid recognition sequence corresponding to the restriction enzyme site during plasmid construction. The commonly used proteases and their corresponding restriction enzyme sites are shown in the following table：