CLONEGENE

GST TAGS

GST can be added to a protein of interest to purify it from solution in a process known as a pull-down assay. This is accomplished by inserting the GST DNA coding sequence next to that which codes for the protein of interest. Thus, after transcription and translation, the GST protein and the protein of interest will be expressed together as a fusion protein. Because the GST protein has a strong binding affinity for GSH, beads coated with the compound can be added to the protein mixture; as a result, the protein of interest attached to the GST will stick to the beads, isolating the protein from the rest of those in solution. The beads are recovered and washed with free GSH to detach the protein of interest from the beads, resulting in a purified protein. This technique can be used to elucidate direct protein-protein interactions. A drawback of this assay is that the protein of interest is attached to GST, altering its native state.^[37][38]

A GST-tag is often used to separate and purify proteins that contain the GST-fusion protein. The tag is 220 amino acids (roughly 26 KDa) in size,^[39] which, compared to tags such as the Myc-tag or the FLAG-tag, is quite large. It can be fused to either the N-terminus or C-terminus of a protein. However, many commercially available sources of GST-tagged plasmids include a thrombin domain for cleavage of the GST tag during protein purification.^[37][40]

Mab 5C12 is a high affinity  mouse monoclonal directed against GST. It has been independently  validated by the Online Antibodies  validation  program and can detect at least 1ng of GST protein. Its high affinity makes it  the antibody of choice for ChIP studies