O-Glycoprotease
O-Glycoprotease
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$242.00 SGD
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$242.00 SGD
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Product Details
Product Details
Product Specification
| Synonyms | O糖蛋白酶 |
| Purity | ≥95% by SDS-PAGE |
| Storage Buffer | 20 mM Tris-HCl，100 mM NaCl ，pH 7.5 |
| Stability & Storage |
This product should be stored at -20℃ and can be stored for at least 12 months. Avoid repeated freezing and thawing. |
Background
O-Glycoproteinase is expressed in E. coli and is a highly specific glycoproteinase that can specifically recognize serine or threonine residues of mucin-type O-linked sugars (whether sialylated or not) and cleave them at their N-termini. It can produce O-glycopeptides suitable for mass spectrometry analysis. The enzyme contains a 6X His-Tag, which can be easily removed after the reaction.
Components
Components |
Size 1 |
Size 2 |
O-glycoprotease |
20U |
100U |
1M Tris-HCl（pH=8.0） |
1.0ml |
1.0ml×5 |
1M DTT |
0.1ml |
0.2ml |
1% SDS |
0.2ml |
1.0ml |
500mM 碘乙酸 |
0.1ml |
0.5ml |
Protocol
Instructions (for reference only)
- Mix 10µg of glycoprotein with 20 mM Tris-HCl pH 8.0, 40 mM DTT, 0.1% SDS solution and make the total volume 50µl.
- Heat at 95°C for 10 minutes.
- Cool to room temperature.
- Add 3µl 500mM iodoacetic acid solution.
- Leave at room temperature for 30 minutes.
- Transfer the solution to an ultrafiltration centrifuge tube.
- Centrifuge at 12000 x g for 4 minutes and discard the flow-through.
- Add 400µl 20mM Tris-HCl, pH 8.0 solution.
- Centrifuge at 12000 x g for 4 minutes and discard the flow-through. Add 400µl 20mM Tris-HCl, pH 8.0 solution and repeat once.
- Add 20µl 20mM Tris-HCl, pH 8.0 solution.
- Add 1µl O-glycoproteinase and mix gently.
- Incubate at 37˚C for 2-18 hours.
- Centrifuge at 12000 x g for 4 minutes and collect the supernatant.
Unit Definition
One unit is the amount of enzyme required to cleave more than 90% of a 2 µM O-glycopeptide in 20 mM Tris-HCl, pH 8.0 in a 20 μL reaction at 37°C for 2 hours.
