Ludger Endo F1 enzyme structure

    Endoglycosidase F1


    E-EF01

    Endo F1 cleaves high mannose and some hybrid type N-glycans from peptides and proteins.

      View product documentation: Specsheet / CofA / MSDS

    • Part Number
    • E-EF01
    • Amount of Enzyme
    • 1 U / 60 µL








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









    Product Specifications:


    Endo F1 cleaves Asparagine-linked high mannose and some hybrid oligosaccharides. Core fucosylation reduces the activity by 50 fold. Endoglycosidase F1 will hydrolyze sulfate containing high-mannose chains. 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.

    Additional Endo F Products:
    Endoglycosidase F2 releases biantennary and high mannose glycans (at a 40X reduced rate)
    Endoglycosidase F3 will release triantennarry and fucosylated biantennary N-glycans

    Source: Recombinant Elizabethkingia miricola in E. Coli

    EC: 3.2.1.96

    Alternative Names: Endo F1, Endoglycosidase F1, endo-β-N-acetylglucosaminidase F

    Endo F1 Specificity: Cleaves all asparagine-linked high mannose and some hybrid oligosaccharides

    Contents:
    Ludger Endoglycosidase F1 - Kit contents
    60 µL aliquot of enzyme (1 U) in 20 mM Tris-HCl, pH 7.5
    5x Reaction Buffer – 250 mM sodium phosphate, pH 5.5

    Specific Activity: >16 U/mg
    Activity: >17 U/mL

    Molecular weight: 32,000 daltons

    Suggested usage:
    1. Add up to 200 µg of glycoprotein to an Eppendorf tube. Adjust to 38 µL final volume with de-ionized water.
    2. Add 10 µL 5x Reaction Buffer 5.5
    3. Add 2.0 µL of Endo F1. Incubate 1 hour at 37°C.

    Specific Activity:
    Defined as the amount of enzyme required to catalyze the release of N-linked oligosaccharides from 1 micromole of denatured Ribonuclease B (RNase B) in 1 minute at 37°C, pH 5.5. Cleavage is monitored by SDS-PAGE (cleaved RNase B migrates faster).

    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 F1 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-EF02
  • E-EF03
  • E-EH02
  • E-G001
  • KE-DG01
  • KE-DGMX
  • PNGase F (Peptide N glycosidase F)
  • Endoglycosidase F2
  • Endoglycosidase F3
  • Endoglycosidase H
  • 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. Maley P., R. B. Trimble, A. L. Tarentino and T. H. Plummer Jr. Characterization of glycoproteins and their associated oligosaccharides through the use of endoglycosidases. Anal Biochem 180:195-204 (1989).

2. Plummer, T. H. Jr, A. W. Phelan and A. L. Tarentino. Porcine fibrinogen glycopeptides: substrates for detecting endo-N-acetylglucosaminidases F2 and F3. Anal Biochem 235:98-101 (1996).

3. Reddy A., B. G. Grimwood, T. H. Plummer Jr and A. L. Tarentino. High- level expression of the Endo-beta-N- acetylglucosaminidase F2 gene in E.coli: one step purification to homogeneity. Glycobiology 8:633-636 (1998).

4. Tarentino, A. L., C. M. Gomez and T. H. Plummer Jr. Deglycosylation of Asparagine-Linked Glycans by Peptide:N-Glycosidase F. Biochemistry 24:4665-4671 (1985).

5. Tarentino A. L., G. Quinones, W. P. Schrader, L. M. Changchien and T. H. Plummer Jr. Multiple endoglycosidase (Endo) F activities expressed by Flavobacterium meningosepticum. Endo F1: molecular cloning, primary sequence, and structural relationship to Endo H. J Biol Chem 267:3868-3872 (1992).

6. Tarentino A. L., G. Quinones, L. M. Changchien, and T. H. Plummer Jr. Multiple endoglycosidase F activities expressed by Flavobacterium meningosepticum endoglycosidases F2 and F3: Molecular cloning, primary sequence, and enzyme expression. J Biol Chem 268(13):9702-9708 (1993).

7. Tarentino A. L. and T. H. Plummer Jr. Substrate specificity of Flavobacterium meningosepticum: Endo F2 and endo F3: purity is the name of the game. Glycobiology 4:771-773 (1994).

8. Tarentino, A. L. and T. H. Plummer Jr. Enzymatic deglycosylation of asparagine- linked glycans: purification, properties and specificity of oligosaccharide- cleaving enzymes from Flavobacterium meningosepticum. Methods in Enzymology 230:44-57 (1994).

9. Tarentino A. L., G. Quinones and T. H. Plummer Jr. Overexpression and purification of non-glycosylated recombinant endo-beta-N- acetylglucosaminidase F3. Glycobiology 5:599-601 (1995).

10. Trimble, R. B. and A. L. Tarentino. Identification of Distinct Endoglycosidase (Endo) Activities in Flavobacterium meningosepticum: Endo F1, Endo F2 and Endo F3. J. Biol Chem 266:1646-1651 (1991).