Active Recombinant Enterococcus faecalis O-Glycosidase, His tagged

• Cat.No.: O-Glycosidase-14E
• Product Overview: O-Glycosidase, also known as endo-α-N-acetylgalactosaminidase, is derived from Enterococcus faecalis and is recombinantly expressed in E.coli. RNase Activity: Negative

Specification

• Species: Enterococcus faecalis
• Source: E.coli
• Tag: His
• Description: After the terminal sialic acid of glycoproteins is removed with neuraminidase, O-Glycosidase can catalyze the removal of O-linked disaccharides with core 1 and core 3 structures from glycoproteins. The storage solution of this product does not contain glycerol, so it has good compatibility with downstream mass spectrometry and HPLC. Moreover, it contains a His tag, which allows for convenient removal by chromatography.
• Purity: ≥ 95% by SDS-PAGE
• Advantages: 1. High purity: It is free from contamination by other proteases and has no other endo- and exo-glycosidase activities, with a purity of ≥ 95%; 2. High stability: Each batch of O-glycosidase undergoes strict quality control to ensure batch-to-batch stability; 3. Compatible with HPLC/ MS: It does not contain glycerol, which helps to achieve optimal results in HPLC and mass spectrometry analyses; 4. Easy removal: It contains a 6×His-Tag and can be easily removed from the reaction system using nickel-affinity resin.
• Protein Content: ≥ 40000 U/μL
• Unit definition: One unit (1 U) is defined as the amount of the enzyme required to remove 0.68 nmol of O-linked glycans from 5 mg of non-denatured fetuin treated with sialidase in a 100 μL reaction system at 37 centigrade within 1 hour.
• Usage: 1. Denaturing enzymatic digestion. Dissolve 10-20 μg of glycoprotein in deionized water. Add 1 μL of 10× glycoprotein denaturation buffer and adjust the volume to 10 μL with deionized water. Incubate at 100 centigrade for 10 min. Transfer to ice and centrifuge for 10 s. Add 2 μL of 10× O-Glycosidase digestion reaction buffer, 2 μL of 10% NP-40, 2 μL of neuraminidase (50 U/μL), 1-4 μL of O-glycosidase, and adjust the volume to 20 μL with water. Mix well. Incubate at 37 centigrade for 1-4 h. Perform SDS-PAGE or HPLC analysis. 2. Non-denaturing enzymatic digestion Dissolve 10-20 μg of glycoprotein in deionized water. Add 2 μL of 10× O-Glycosidase digestion reaction buffer, 2 μL of neuraminidase (50 U/μL), 2-5 μL of O-glycosidase, and adjust the volume to 20 μL with deionized water. Incubate at 37 centigrade for 2-18 h. If the digestion effect is poor, appropriately increase the amount of enzyme and extend the digestion time. Perform SDS-PAGE or HPLC analysis.
• Applications: 1. Determination of O-glycosylation sites 2. Determination of O-glycan structures
• Note: 1. Try to avoid freeze-thaw cycles of this product after receipt; 2. Please wear lab coat and disposable gloves when using; 3. This product should not be used directly for clinical diagnosis and treatment.
• Stability: This product should be stored at -20 centigrade and can be stored for at least 12 months.
• Storage: Store at -20 centigrade.
• Storage Buffer: 20 mM Tris-HCl, 50 mM NaCl, 1 mM EDTA, pH 7.5
• Shipping: Dry Ice