Glutathione S-transferase Tag
Glutathione S-transferase Tag - Related Videos
14:57
Spectrophotometric Screening for Potential Inhibitors of Cytosolic Glutathione S-Transferases
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Cited by 14 •
2020
Glutathione S-transferases (GSTs) are detoxification enzymes involved in the metabolism of numerous chemotherapeutic drugs. Overexpression of GSTs is correlated with cancer chemotherapy resistance. One way to counter this phenotype is to use inhibitors. This protocol describes a method using a spectrophotometric assay to screen for potential GST inhibitors.
08:10
Measuring Glutathione-induced Feeding Response in Hydra
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Cited by 9 •
2014
Here we describe a simple assay for the quantification of the feeding response in hydra induced by the reduced form of glutathione. This assay relies on measuring the distance between the apical end of the tentacle and mouth of hydra.
07:46
An In Vitro Assay to Detect tRNA-Isopentenyl Transferase Activity
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Cited by 3 •
2018
Here, we describe a protocol for the biochemical characterization of the yeast RNA-modifying enzyme, Mod5, and discuss how this protocol could be applied to other RNA-modifying enzymes.
11:26
High-throughput Gene Tagging in Trypanosoma brucei
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Cited by 21 •
2016
Addition of a tag to a protein is a powerful way of gaining insight into its function. Here, we describe a protocol to endogenously tag hundreds of Trypanosoma brucei proteins in parallel such that genome scale tagging is achievable.
05:32
Terminal Transferase-Mediated dUTP Nick End-Labeling—TUNEL Assay: An In Situ Method to Detect DNA Fragmentation in Apoptotic Cells
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2025
In this video, the TUNEL assay is performed to detect the presence of apoptotic cells in a sample. The study involves using a special DNA polymerase - terminal deoxynucleotidyl transferase - that catalyzes the addition of labeled dUTPs to the 3' terminus of fragmented DNA, which can be observed and identified under a fluorescence microscope.
FAQs
The GST portion of a fusion protein binds glutathione on a conjugated resin, creating a selective capture step during purification. After unbound lysate components are removed, the retained fusion can be released under controlled conditions. This affinity interaction is the central mechanism that enriches the recombinant target from a complex biological mixture.
GST can improve the soluble expression of some recombinant target proteins, although the effect depends on the target. Greater solubility can make the fusion protein more accessible for affinity purification from a cell lysate and may support subsequent detection or biochemical analysis. The tag therefore contributes not only to capture, but also to the handling of certain expressed proteins.
Enzymatic cleavage allows the GST portion to be separated from the target protein after the fusion has been purified. Removing the tag can be useful when downstream experiments require the target in a less modified form, such as structural analysis or biochemical assays. Cleavage adds a controlled processing step between affinity purification and later characterization.
A typical workflow begins with a cell lysate containing the recombinant fusion protein. The lysate is applied to glutathione-conjugated resin so the GST portion can bind, while other components remain unretained. The captured fusion is then released under controlled conditions, and enzymatic cleavage can be used when the target must be separated from its tag.
GST-tagged proteins support several protein-science applications beyond isolation. Their capture and detection can help researchers obtain material for interaction studies, structural analysis, and biochemical assays. In interaction experiments, the tagged protein can serve as an identifiable or recoverable fusion partner, while purified material provides a defined input for examining structure or biochemical behavior.
In molecular biology, the tag connects recombinant gene expression with practical protein analysis. A genetically attached GST sequence can support detection, affinity-based enrichment from cell lysates, and assessment of soluble expression. The resulting fusion protein can then be used in experiments that examine molecular interactions, structure, or biochemical activity, linking expression methods to downstream protein characterization.