Frequently Asked Questions
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When and why should I use the OpeRATOR elution procedure?
The OpeRATOR enzyme digests O-glycosylated proteins N-terminally to the serine (S) and threonine (T) glycosylation sites. OpeRATOR therefore facilitates mapping of O-glycosylation sites by LC-MS and as the digested products no longer bind to the GlycOCATCH resin, elution and sample preparation can be achieved in a single step. However, this requires the O-glycosylation sites to be spaced closely together for the OpeRATOR digestion to yield peptides of a size suitable for analysis by LC-MS/MS.
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Is it possible to digest the glycoprotein with OpeRATOR first and then enrich the O-glycopeptides on GlycOCATCH?
No, when the glycoprotein is digested with OpeRATOR, the binding capacity of GlycOCATCH is lost.
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Can I load a digested protein solution directly onto the GlycOCATCH column?
Yes. IMPORTANT! Before you load your sample make sure that the protease in solution is inactivated and that the pH, buffer, chaotrope and NaCl concentrations are within the recommended ranges according to the instructions.
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Does GlycOCATCH bind all O-glycosylated proteins and peptides?
No, GlycOCATCH is designed to bind specifically to mucin type (core 1) O-glycosylated proteins and peptides. The performance is significantly enhanced by removal of sialic acids using SialEXO.
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