His-tag

Definition

A His-tag, or Histidine-tag, is a protein purification and detection tool that involves adding a short peptide sequence, typically consisting of six to ten consecutive histidine residues, to a recombinant protein. This tag allows for the efficient purification and identification of the target protein.

5 Must Know Facts For Your Next Test

1. The His-tag is typically added to the N-terminus or C-terminus of a recombinant protein during the genetic engineering process.
2. The histidine residues in the His-tag have a high affinity for metal ions, such as nickel or cobalt, which are immobilized on a solid support in affinity chromatography columns.
3. The His-tagged protein can be selectively bound to the metal-containing resin, allowing for the separation and purification of the target protein from other cellular components.
4. The His-tag is often used in pharmaceutical applications to purify recombinant proteins, such as enzymes, antibodies, or therapeutic proteins, for further characterization and use.
5. The small size of the His-tag (typically 6-10 amino acids) minimizes the risk of interfering with the structure and function of the target protein.

Review Questions

• Explain how the His-tag is utilized in the purification of recombinant proteins.
  • The His-tag, a short peptide sequence consisting of consecutive histidine residues, is genetically engineered onto a recombinant protein during the expression process. This His-tag has a high affinity for metal ions, such as nickel or cobalt, which are immobilized on a solid support in affinity chromatography columns. The His-tagged protein can then be selectively bound to the metal-containing resin, allowing for the separation and purification of the target protein from other cellular components. This method of affinity chromatography enables the efficient isolation and concentration of the desired recombinant protein for further analysis and applications.
• Discuss the advantages of using the His-tag system in pharmaceutical applications of genetic engineering.
  • The His-tag system offers several advantages in the pharmaceutical applications of genetic engineering. Firstly, the small size of the His-tag (typically 6-10 amino acids) minimizes the risk of interfering with the structure and function of the target protein, which is crucial for maintaining the therapeutic properties of recombinant proteins. Secondly, the high affinity of the His-tag for metal ions allows for the efficient purification of the target protein from complex mixtures, such as cell lysates, ensuring a high degree of purity for downstream applications. Additionally, the His-tag can be used as a detection tool, facilitating the identification and tracking of the recombinant protein during the various stages of pharmaceutical development and production.
• Analyze the role of the His-tag in the context of whole genome methods and how it contributes to the advancement of genetic engineering techniques.
  • The His-tag plays a crucial role in the context of whole genome methods and the advancement of genetic engineering techniques. In whole genome studies, the His-tag can be used to facilitate the purification and analysis of proteins encoded by genes identified through genome sequencing and annotation. By allowing for the efficient isolation of target proteins, the His-tag enables researchers to study the structure, function, and interactions of these proteins, which is essential for understanding the complex mechanisms underlying biological processes at the genomic level. Furthermore, the His-tag system is widely adopted in genetic engineering workflows, as it provides a reliable and versatile tool for the purification and characterization of recombinant proteins, which are central to many pharmaceutical and biotechnological applications. The ease of use and the scalability of the His-tag system have contributed to the rapid progress in genetic engineering, empowering researchers to explore and exploit the vast potential of the genome for various scientific and industrial purposes.

Related terms

Affinity Chromatography: A separation technique that utilizes the specific interaction between a target molecule and a ligand immobilized on a solid support to purify the molecule of interest.

Recombinant Protein: A protein that is produced by inserting a gene encoding the desired protein into a host organism, such as bacteria or yeast, which then expresses the protein.

Protein Purification: The process of isolating a specific protein from a complex mixture, such as a cell lysate, to obtain a pure and concentrated sample for further analysis or applications.