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OpeRATOR is an O-protease digesting O-glycosylated proteins, with mucin-type O-glycosylation, N-terminally of the S/T glycosylation site. The presence of O-glycans is required for OpeRATOR to exert its enzymatic activity.
OpeRATOR is an endoprotease that catalyzes the hydrolysis of peptide bonds in native mucin-type O-glycosylated proteins in direct proximity of the O-glycans. With high specificity, OpeRATOR digests proteins at the N-terminus of O-glycans at serine or threonine. O-glycans of mucin-type are required for OpeRATOR activity and the enzyme will not digest glycoproteins with only N-linked glycans. OpeRATOR is active on O-glycan proteins with or without sialic acids, although the activity is higher on O-glycan proteins where sialic acids have been removed. OpeRATOR can be used together with SialEXO (sialidase), which is included in the box.
OpeRATOR is derived from Akkermansia muciniphila and expressed in E. coli. The enzyme contains a His-tag and the molecular weight is 42 kDa. SialEXO is derived from Akkermansia muciniphila and expressed in E. coli. The enzymes in the SialEXO also contain a His-tag and the molecular weights of these components are 43 kDa and 66 kDa, respectively.
One unit OpeRATOR digests ≥ 90% of 1 µg glycoprotein (TNFαR) when incubated together with one unit of SialEXO in 20 mM Tris pH 6.8 at 37 °C for 2 h.
Content and Storage
The OpeRATOR box includes:
1 vial of OpeRATOR enzyme (G1-OP1-020) supplied lyophilized in TBS pH 7.6, with no preservatives added.
1 vial of SialEXO (G1-SM1-020) supplied lyophilized in TBS pH 7.6, with no preservatives added.
The vials in the OpeRATOR box is shipped cold and should be stored at –20 °C upon arrival. After reconstitution the enzymes are stable for 1 month at +4-8 °C.
Digests native mucin-type O-glycosylated proteins
2h to overnight (16-18h) reaction
Desialylation using SialEXO® (included) increases performance
N-terminally of O-glycosylated serine and threonine residues
For more information about OpeRATOR® Smart Enzymes