Ludger Endo H enzyme structure

    Endoglycosidase H


    E-EH02

    Endo H cleaves asparagine-linked hybrid or high mannose oligosaccharides but not complex oligosaccharides.

      View product documentation: Specsheet / CofA / MSDS

    • Part Number
    • E-EH02
    • Amount of Enzyme
    • 0.3 U / 60 µL








    Kit includes enzyme plus reaction buffers.
    Sufficient for up to 60 reactions.








    Product Specifications:


    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.





Companion Products:

Process controls


Glycan clean-up - removal of proteins and enzymes from glycan mixtures following endoglycosidase treatment

  • LC-PBM-96
  • LudgerClean Protein Binding Membrane (96 samples)

Labelling and derivatisation of released glycans - View our Glycan Labelling Summary Table

  • LT-KAB-A2
  • LT-KAB-VP24
  • LT-KAB-VP96
  • LT-KPROC-24
  • LT-KPROC-96
  • LT-KPROC-VP24
  • LT-KAA-A2
  • LT-KAA-VP24
  • LT-PERMET-96
  • LT-PERMET-VP96
  • LudgerTag 2-AB Glycan Labelling Kit, sodium cyanoborohydride (24 samples)
  • LudgerTag 2-AB Glycan Labelling Kit, picoline borane (24 samples)
  • LudgerTag 2-AB Glycan Labelling Kit, picoline borane (96 samples)
  • LudgerTag Procainamide Glycan Labelling Kit, sodium cyanoborohydride (24 samples)
  • LudgerTag Procainamide Glycan Labelling Kit, sodium cyanoborohydride (96 samples)
  • LudgerTag Procainamide Glycan Labelling Kit, picoline borane (24 samples)
  • LudgerTag 2-AA Glycan Labelling Kit, sodium cyanoborohydride (24 samples)
  • LudgerTag 2-AA Glycan Labelling Kit, picoline borane (24 samples)
  • LudgerTag Permethylation Kit (96 samples)
  • LudgerTag Permethylation Kit, without methyl iodide (96 samples)

Alternative enzymes

  • LZ-rPNGaseF-kit
  • E-EF01
  • E-EF02
  • E-EF03
  • E-G001
  • KE-DG01
  • KE-DGMX
  • PNGase F (Peptide N glycosidase F)
  • Endoglycosidase F1
  • Endoglycosidase F2
  • Endoglycosidase F3
  • O-glycosidase
  • Enzymatic CarboRelease Kit - contains enzymes necessary for removing N-linked and O-linked oligosaccharides
  • Enzymatic DeGlycoMx Kit - contains enzyme mix for removing N-linked and O-linked oligosaccharides






References:

1. Robbins P. W., D. F. Wirth and C. J. Hering. Expression of the Streptomyces enzyme endoglycosidase H in Escherichia coli. Biol Chem 256:10640-10644 (1981).

2. Robbins P. W., R. B. Trimble, D. F. Wirth, C. Hering, F. Maley , G. F. Maley, R. Das, B. W. Gibson, N. Royal and K. Biemann. Primary structure of the Streptomyces enzyme endo-beta-N-acetylglucosaminidase H. J Biol Chem 259:7577-7583 (1984).

3. Trimble R. B., A. L. Tarentino , G. E Aumick and F. Maley. Endo-beta-N-acetylglucosaminidase L from Streptomyces plicatus. Methods Enzymol 83:603-610 (1982).

4. Trimble R. B. and F. Maley. Optimizing hydrolysis of N-linked high-mannose oligosaccharides by endo-beta-N-acetylglucosaminidase H. Anal Biochem 141:515-522 (1984).

5. Trimble R. B., R. J. Trumbly and F. Maley. Endo-beta-N-acetylglucosaminidase H from Streptomyces plicatus. Methods Enzymol 138:763-770 (1987).

6. Trumbly R. J., P. W. Robbins, M. Belfort, F. D. Ziegler, F. Maley and R. B. Trimble. Amplified expression of Streptomyces endo-beta-N-acetylglucosaminidase H in Escherichia coli and characterization of the enzyme product. J Biol Chem 260:5683-5690 (1985).