GST-tag

Definition

A GST-tag is a fusion protein that combines a protein of interest with the glutathione S-transferase (GST) protein. This tag is commonly used in molecular biology and biochemistry to facilitate the purification and detection of recombinant proteins.

5 Must Know Facts For Your Next Test

1. The GST-tag allows for the easy purification of the recombinant protein using affinity chromatography with a glutathione-agarose resin.
2. The GST-tag can also be used to enhance the solubility of the target protein, making it easier to express and purify.
3. The GST-tag can be cleaved off the target protein using a specific protease, such as thrombin or PreScission protease, to obtain the pure, tag-free protein.
4. GST-tagged proteins can be detected and quantified using antibodies that recognize the GST tag.
5. The GST-tag is commonly used in the pharmaceutical applications of genetic engineering, such as the production of therapeutic proteins and the screening of drug candidates.

Review Questions

• Explain how the GST-tag is used in the purification of recombinant proteins.
  • The GST-tag is used to facilitate the purification of recombinant proteins through affinity chromatography. The GST protein has a high affinity for the immobilized ligand glutathione, which is present on a solid support, such as agarose beads. When the GST-tagged recombinant protein is applied to the affinity column, the GST-tag binds to the glutathione, allowing the target protein to be separated from the rest of the cellular components. The purified protein can then be eluted from the column using a buffer containing free glutathione, which competes with the immobilized glutathione for binding to the GST-tag.
• Discuss the role of the GST-tag in the pharmaceutical applications of genetic engineering.
  • The GST-tag is commonly used in the pharmaceutical applications of genetic engineering, particularly in the production of therapeutic proteins and the screening of drug candidates. The ability to easily purify recombinant proteins using the GST-tag is crucial for the large-scale production of therapeutic proteins, such as enzymes, antibodies, or cytokines, which are used to treat various medical conditions. Additionally, the GST-tag can be used to immobilize potential drug candidates on a solid support, allowing for the screening of libraries of compounds for their ability to bind to a target protein of interest, which is an important step in the drug discovery process.
• Analyze how the GST-tag can be used to study protein-protein interactions in the context of whole genome methods.
  • In the context of whole genome methods, the GST-tag can be used to study protein-protein interactions on a large scale. By fusing the GST-tag to a protein of interest, the tagged protein can be used as 'bait' to pull down interacting 'prey' proteins from a complex mixture, such as a cell lysate. The GST-tag allows for the efficient purification of the bait protein and any associated prey proteins, which can then be identified using mass spectrometry or other analytical techniques. This approach, known as affinity purification-mass spectrometry (AP-MS), is a powerful tool for mapping protein-protein interaction networks and understanding the functional relationships between proteins in a whole-genome context.

Related terms

Fusion Protein: A recombinant protein that consists of two or more proteins or protein domains joined together, often used to study protein-protein interactions or facilitate protein purification.

Affinity Chromatography: A separation technique that uses the specific interaction between a target molecule and a ligand immobilized on a solid support to purify the target from a complex mixture.

Recombinant Protein: A protein that is produced by inserting a gene encoding the protein of interest into a host organism, such as bacteria or yeast, and then expressing and purifying the resulting protein.