O-glycan-specific Protein Digestion
ImpaRATOR is an O-glycan-dependent protease that digests proteins carrying mucin-type
O-glycans, including sialylated species, N-terminally of glycosylated Ser and Thr residues.
The enzyme generates glycopeptides carrying O-glycans, which enables O-glycan profiling, site occupancy determination and O-glycopeptide mapping as well as middle-level approaches using LC-MS analysis.
- Broad specificity - accepts a wide range of O-glycan variants, including sialylated species
- Reliable O-glycan-specific protease – digests N-terminally of Ser/Thr glycosylation sites
- Robust digestion for enhanced characterization of complex biopharmaceuticals
Download product folderO-glycosylated
Ser and Thr
ImpaRATOR™ Lyophilized
Lyophilized enzyme for digestion of mucin-type O-glycoproteins and peptides, including sialylated O-glycan speciesAbout ImpaRATOR™
ImpaRATOR is an O-glycan-dependent protease that catalyzes the hydrolysis of the peptide bond adjacent to glycosylated serine or threonine residues in glycoproteins and glycopeptides. It cleaves N-terminally of serine or threonine residues modified with mucin-type O-glycans, including sialylated species.
Mucin-type O-glycans are required for ImpaRATOR activity and the enzyme will not digest unmodified serine or threonine residues, or at N-glycosylation sites of glycoproteins. The enzyme accepts a broad range of O-glycan structures, including sialylated core 1 and core 2 structures as well as the Tn antigen.
ImpaRATOR is derived from Pseudomonas aeruginosa and expressed in E. coli. The enzyme contains a His-tag and has a molecular weight of 97 kDa.
ImpaRATOR™ Lyophilized
Lyophilized enzyme for digestion of mucin-type O-glycoproteins and peptides, including sialylated O-glycan species.
ImpaRATOR Lyophilized is available as a lyophilized powder in 2000 unit vials for digestion of 2 mg O-glycosylated protein.
- Flexible format for method development
- Possible to combine enzymes for improved efficiency
- Amenable to automated workflows
- Ready to use – just add water!
Available Products
For information on how to order, visit Place an Order or contact us directly at order@genovis.com.
Comprehensive Analysis ofUnit Definition
One unit ImpaRATOR Lyophilized digests ≥ 95% of 30 µg etanercept at least at one site when incubated in TBS (50 mM Tris-HCl, 150 mM NaCl), pH 7.6 at 37°C for 30 minutes. For O-glycopeptide mapping applications, one unit ImpaRATOR Lyophilized digests 1 µg O-glycosylated protein when incubated in TBS, pH 7.6 at 37°C for 2 hours.
Content and Storage
ImpaRATOR Lyophilized is supplied lyophilized in TBS (50 mM Tris-HCl, 150 mM NaCl), pH 7.6, with no preservatives added.
ImpaRATOR Lyophilized is shipped at ambient temperature, and should be stored at -20°C upon arrival.
After reconstitution, the ImpaRATOR enzyme is stable for at least 1 month at +4-8°C.
Instructions
Product Specification
Safety Data Sheet
Certificate of Analysis
FAQ and Support
Popular FAQ
Insufficient digestion during non-denaturing conditions can be caused by O-glycosylation sites within the glycoprotein inaccessible to the enzyme. In such cases, we recommend trying the following workflow: reduction, denaturation, carboxymethylation, buffer-exchange, and then digestion with ImpaRATOR. If detergent is added, make sure to include a detergent removal step prior ImpaRATOR digestion.
ImpaRATOR has a broad activity towards different O-glycan structures; however, the enzyme has limited activity towards sites with two adjacent O-glycosylated Ser/Thr residues. For complete information about O-glycan sites in glycoprotein substrates containing several sites with two adjacent O-glycosylated Ser/Thr residues, OpeRATOR may be a better option.
ImpaRATOR is a metalloprotease and thereby sensitive to chelating agents such as EDTA. Concentrations > 1 mM EDTA results in complete inhibition of the enzyme. In addition, the ImpaRATOR activity is inhibited by reducing agents and detergents.
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