Recombinant proteins have shown great differences in solubility, stability, and functionality during their expression and purification, which makes it difficult to perform large-scale protein production. Addition of fusion tags is the best way to effectively improve the expression level, solubility, and stability of recombinant proteins, especially for proteins that is difficult to express.
The most frequently used fusion tag to collect large amounts of high-quality protein is the His-tag. It usually composes of 6-14 histidines, to which the N- or C-terminal of the target protein is typically fused. The His-tag contains imidazole side chains for irreversibly engagement of coordinative bonds to Ni2+, Co2+, or Zn2+. Ni2+ is preferred because of the greatest affinity and selectivity for His-tags. His-tagged proteins prefer to form a considerably stronger bond to the Ni-NTA resin than any endogenous protein that contains histidine. How many histidines bind to the matrix directly affect the relative binding strength.
Advantages of His-tag:
1. Easy control of the number of the histidine in a His-tag. The longer His-tag is, the better binding and separation it delivers;
2. His-tags are much smaller than other fusion tags;
3. Purification can be performed under native and denaturing conditions;
4. Increase the solubility of target proteins;
5. Rarely interfere with function of target proteins.
His-tagged Protein Purification Troubleshooting
His-tagged proteins do not bind to Ni-NTA
His-tag is not present
His-tag is inaccessible
His-tagged proteins precipitation
Creative Biostructure is happy to accelerate your His-tagged protein research project from plasmid construction to high-quality recombinant protein production. Please see the table below for more information and feel free to Contact Us.