Endo H cleaves Asparagine-linked hybrid or high mannose oligosaccharides, but not complex oligosaccharides. It cleaves between the two N-acetylglucosamine residues in the diacetylchitobiose core of the oligosaccharide, generating a truncated sugar molecule with one N-acetylglucosamine residue remaining on the asparagine. In contrast, PNGase F removes the oligosaccharide intact. Detergent and heat denaturation may increase the rate of cleavage for some glycoproteins.

Source: recombinant from Streptomyces plicatus in E.Coli

EC: 3.2.1.96

Alternative Names: Endo H, endo-β-N-acetylglucosaminidase H, Endoglycosidase H

Specific Activity: One unit of Endo H activity is defined as the amount of enzyme required to catalyze the release of N-linked oligosaccharides from 1 µmole of denatured Ribonuclease B. Cleavage is monitored by SDS-PAGE (cleaved Ribonuclease B migrates faster).

Contents:
60 µL aliquot of enzyme (300 mU) in 20 mM Tris-HCl, 25 mM NaCl, 1 mM EDTA (pH 7.5).
200 µL 5x Reaction Buffer 5.5 (250 mM sodium phosphate, pH 5.5)
Denaturation Solution - 2% SDS, 1 M Beta-mercaptoethanol

Specific Activity: >40 U/mg

Activity: >5 U/mL

Molecular weight: 29,000 daltons

pH range: 5-6, optimum 5.5

Suggested usage:
1. Add up to 200 µg of glycoprotein to Eppendorf tube. Adjust to 37.5 µL final volume with deionized water.
2. Add 10 µL 5X Endo H Buffer and 2.5 µL of Denaturation Solution (SDS/-ME). Heat at 100°C for 5 minutes.
3. Cool, and then add 2.0 µL of Endo H to the reaction. Incubate 3 hours at 37°C.

Storage: Store enzyme at 4°C.

Stability: Stable at least 12 months when stored properly. Several days exposure to ambient tempertures will not reduce activity.

Purity: Endoglycosidase H is tested for contaminating protease as follows; 10 µg of denatured BSA is incubated for 24 hours at 37°C with 2 µL of enzyme. SDS-PAGE analysis of the treated BSA shows no evidence of degradation.

The production host strain has been extensively tested and does not produce any detectable glycosidases.