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.
Coupled Protein Expression & Purification includes these areas of focus:
FAQs for Coupled Protein Expression & Purification
Protocols for Coupled Protein Expression & Purification
Application Notes Coupled Protein Expression & Purification
- Co-expression of Multiple Proteins in Kluyveromyces lactis
- Intein-Mediated Protein Ligation (IPL) and Labeling with the IMPACT™ Kit
- Protein Expression with T7 Express Strains
- Transformation of SHuffle® Competent Cell Strains
- Use of the PURExpress® in vitro Protein Synthesis Kit, Disulfide Bond Enhancer and SHuffle® Competent E. coli for heterologous in vitro and in vivo cellulase expression.
Bypassing Common Obstacles in Protein Expression
The Next Generation of Cell-free Protein Synthesis
Why Choose the K. lactis Protein Expression Kit?
Review the advantages of the K. lactis Protein Expression Kit for rapid, high yield protein expression in yeast.
Protein Expression & Purification Brochure
The Protein Expression and Purification brochure provides information on the advanced tools for protein expression and purification offered by NEB.
- IMPACT™ Vectors and Applications
- Protein Expression and Purification Selection Chart
Other Tools & Resources
Overview of Secretion in K. lactis
The pKLAC2 Expression Vector
Expression of MBP with K. lactis
pKLAC2 Multiple Cloning Site
Schematic Illustration of Purification Using pMAL™ System
Protein Expression Using pMAL™
Schematic Illustration of the IMPACT® System
Expression and Purification of E. coli Maltose-Binding Protein (MBP) Using the IMPACT System
NEB has a long history in recombinant protein expression and has developed a wide array of solutions for proteins that are difficult to express.