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GST-tagged Proteins

The Glutathione S-transferase (GST) fusion system is a comprehensive system to express, purify, and detect GST-tagged proteins. The GST affinity tag allows protein purification under mild conditions and easy cleavage, keeping the proteins in their native structure and function.Three major parts comprise this system: pGEX plasmid expression vectors, GST-tagged protein purification, and GST-tagged protein detection.

Advantage of GST fusion system:
1. All pGEX vectors provide a tac promoter to express proteins at high-level and with chemical induction;
2. Purify active proteins under mild and nondenaturing conditions;
3. Suitable for one-step, high-yield affinity chromatography purification based on glutathione sepharose media;
4. Easy cleavage of target protein from fusion protein with multiple proteases such as Thrombin or Factor Xa;
5. Convenient detection of GST fusion protein with antibodies.

GST-Tag
GST is an antioxidant enzyme composed of 211 amino acids (MW, around 26 kDa). It is water soluble and involves of the primary cellular defense mechanismas a dimer in all aerobic organisms. DNA sequence of GST is frequently applied in plasmid construction. The resulting expression vector is used to produce GST-tagged fusion protein, and GST tag enhance the expression level and solubility of recombinant proteins.

GST-tagged Protein Detection
1. GST 96-Well Detection Module; 
This module allows to determine GST fusion proteins in various sample rapidly and sensitively.
2. Anti-GST Antibody;
This polyclonal antibody isolated from the sera of goats can be used to detect recombinant GST-tagged proteins with high sensitivity and specificity.
3. GST Detection Module;
This module permits sensitive determination of GST-tagged proteins based on a biochemical assay or an immunoassay in which glutathione and CDNB used as substrates for GST.

GST-tagged Protein Purification Troubleshooting

Problem Cause Solution
Low protein yield Problems with vector construction Ensure that protein and tag are in frame
Low protein expression Optimize expression conditions
Inclusion bodies Lower the growth temperature
Insufficient extraction Check lysozyme and sonication conditions
Poor binding efficiency Fusion protein is not well concentrated Concentrate the sample
Flow rate is too high Decrease flow rate
Reducing agent missing Add DTT (5 mM) to the lysis buffer
Severe sonication Use milder sonication conditions
Inefficient elution Flow rate is too high Decrease flow rate
Low elution volume Increase the volume of Elution Buffer
Elution Buffer stored for too long Prepare Elution Buffer immediately before use
Poor protein purity Insufficient washing Repeat washing steps
Fusion protein degradation Add a protease inhibitor or perform purification on ice
Severe sonication Use milder sonication conditions

Creative Biostructure is pleased to accelerate your GST-tagged protein research project from expression vector construction to high-yield recombinant protein production. Please see the table below for more information and feel free to Contact Us.