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324727 Endoglycosidase F3, Elizabethkingia meningosepticum, Recombinant, E. coli

Endoglycosidase F3, Elizabethkingia meningosepticum, Recombinant, E. coli, cleaves asparagine-linked or free biantennary and triantennary complex, and Man3GlcNAc oligosaccharides from glycoproteins.

MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.

Synonyms: Endo-β-N-acetylglucosaminidase F3, Endo F3

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324727
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      Description
      OverviewRecombinant, Elizabethkingia meningosepticum endoglycosidase F3 expressed in E. coli. Cleaves asparagine-linked or free biantennary and triantennary complex, and Man3GlcNAc oligosaccharides from glycoproteins. This enzyme 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 residue. Core fucosylation increases the activity of Endo F3 up to 40 fold. Exhibits no activity on high mannose and hybrid molecules. Less sensitive to protein conformation than N-Glycosidase F (Cat. No. 362185) and therefore is more suitable for deglycosylation of native proteins.
      Note: 1 mU = 1 milliunit.
      Catalogue Number324727
      Brand Family Calbiochem®
      SynonymsEndo-β-N-acetylglucosaminidase F3, Endo F3
      References
      ReferencesTarentino, A.L., et al. 1995. Glycobiology 5, 599.
      Tarentino, A.L., and Plummer, T.H. 1994. Methods Enzymol. 230, 44.
      Tarentino, A.L., et al. 1993. J. Biol. Chem. 268, 9702.
      Trimble, R.B., and Tarentino, A.L. 1991. J. Biol. Chem. 266, 1646.
      Product Information
      Activity≥5 units/ml
      Unit of DefinitionOne unit is defined as the amount of enzyme that will release N-linked oligosaccharides from 1.0 µmol porcine fibrinogen per min at 37°C, pH 5.5.
      EC number3.2.1.96
      FormLiquid
      FormulationIn 20 mM Tris-HCl, pH 7.5.
      Quality LevelMQ100
      Applications
      Biological Information
      Specific Activity≥30 units/mg protein
      Physicochemical Information
      ContaminantsProteases: none detected
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Storage and Shipping Information
      Ship Code Blue Ice Only
      Toxicity Standard Handling
      Storage +2°C to +8°C
      Do not freeze Yes
      Packaging Information
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Catalogue Number GTIN
      324727-200MIU 04055977216011

      Documentation

      Endoglycosidase F3, Elizabethkingia meningosepticum, Recombinant, E. coli SDS

      Title

      Safety Data Sheet (SDS) 

      Endoglycosidase F3, Elizabethkingia meningosepticum, Recombinant, E. coli Certificates of Analysis

      TitleLot Number
      324727

      References

      Reference overview
      Tarentino, A.L., et al. 1995. Glycobiology 5, 599.
      Tarentino, A.L., and Plummer, T.H. 1994. Methods Enzymol. 230, 44.
      Tarentino, A.L., et al. 1993. J. Biol. Chem. 268, 9702.
      Trimble, R.B., and Tarentino, A.L. 1991. J. Biol. Chem. 266, 1646.
      Data Sheet

      Note that this data sheet is not lot-specific and is representative of the current specifications for this product. Please consult the vial label and the certificate of analysis for information on specific lots. Also note that shipping conditions may differ from storage conditions.

      Revision19-June-2008 RFH
      SynonymsEndo-β-N-acetylglucosaminidase F3, Endo F3
      DescriptionRecombinant, Elizabethkingia meningosepticum endoglycosidase F3 expressed in E. coli. Cleaves asparagine-linked or free biantennary and triantennary complex, and Man3GlcNAc oligosaccharides from glycoproteins. This enzyme 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 residue. Core fucosylation increases the activity of Endo F3 up to 40 fold. Exhibits no activity on high mannose and hybrid molecules. Less sensitive to protein conformation than N-Glycosidase F (Cat. No. 362185) and therefore is more suitable for deglycosylation of native proteins.
      FormLiquid
      FormulationIn 20 mM Tris-HCl, pH 7.5.
      Recommended reaction conditions50 mM sodium acetate buffer, pH 4.5 Endoglycosidase F3, Chryseobacterium meningosepticum, Recombinant, E. coli Note: This protocol is provided only as a general guideline. Researchers should standardize this assay for their own specific needs and should consult published literature. Endoglycosidase F3 cleaves asparagine-linked or free biantennary and triantennary complex oligosaccharides and Man3GlcNAc oligosaccharides from glycoproteins. It cleaves between the two N-acetylglucosamine residues in the diacetylchitobiose core of the oligosaccharide, generating a truncated sugar molecule with one Nacetylglucosamine residue remaining on the asparagine residue. Core fucosylation increases the activity of the Endoglycosidase F3 up to 40-fold. Endoglycosidase F3 is less sensitive to protein conformation than N-Glycosidase F (Cat. No. 362185) and is, therefore, more suitable for deglycosylation of native proteins. Endo F3 is highly sensitive to the presence of detergents, so it is recommended that any residual detergents be removed prior to deglycosylation. Required Reagents • Substrate: porcine fibrinogen (6.1 mg/ml) in 1X assay buffer • Diluted enzyme preparation: dilute the endoglycosidase F3 1:400 in 1X assay buffer • 5X assay buffer: 250 mM sodium acetate, pH 4.5 • Triton® X-100 detergent (Cat. No. 648462): 15% solution • SDS-PAGE system Protocol 1. Add 50 µl (304 µg) of substrate to each of 6 tubes. 2. Add 0, 1, 2, 3, 4, or 5 µl of diluted enzyme to appropriately labeled tubes and incubate 1 h at 37°C. 3. Stop the reaction by heating to 100°C for 5 min. 4. Run 10 µl of each sample on a 10% SDS-PAGE gel. Stain with Coomassie Blue. 5. Note the lowest tube number in which ~50% of the substrate is cleaved. 6. Calculate the activity: Units/ml = µmol/min/ml = (8 X 105) R/M T V R = 152 (µg of fibrinogen cleaved M = 185,000 (formula weight of fibrinogen) T = 60 (time in min) V = volume (µl) of enzyme dilution needed to cleave 50% For example: if 2 µl of diluted enzyme results in 50% cleavage, the activity would be 5.5 U/ml. Assay for Deglycosylation Required Reagents • 5X assay buffer: 250 mM sodium acetate, pH 4.5 • sterile dH2O • glycoprotein sample • Endoglycosidase F3 • SDS-PAGE system Protocol 1. Add up to 200 µg of glycoprotein to an eppendorf tube and adjust the final volume to 37.5 µl with dH2O. 2. Add 10 µl of 5X assay buffer. 3. Add 2 µl of Endoglycosidase F3 to the reaction, mix well, and incubate for 1 h at 37°C. Note: When cleaving non-core fucosylated biantennary and triantennary N-linked oligosaccharides, increase the incubation time to 15 days. Many glycoproteins are contaminated with proteases. We advise adding suitable protease inhibitors if long duration incubations are anticipated. As an alternative to long duration incubations with Endoglycosidase F3 for removal of other classes of N-linked oligosaccharides, we suggest using our Glycoprotein Deglycosylation Kit (Cat. No. 362280). 4. Monitor cleavage by SDS-PAGE. Stain with Coomassie Blue.
      EC number3.2.1.96
      ContaminantsProteases: none detected
      Specific activity≥30 units/mg protein
      Activity≥5 units/ml
      Unit definitionOne unit is defined as the amount of enzyme that will release N-linked oligosaccharides from 1.0 µmol porcine fibrinogen per min at 37°C, pH 5.5.
      Storage +2°C to +8°C
      Do Not Freeze Yes
      Toxicity Standard Handling
      ReferencesTarentino, A.L., et al. 1995. Glycobiology 5, 599.
      Tarentino, A.L., and Plummer, T.H. 1994. Methods Enzymol. 230, 44.
      Tarentino, A.L., et al. 1993. J. Biol. Chem. 268, 9702.
      Trimble, R.B., and Tarentino, A.L. 1991. J. Biol. Chem. 266, 1646.