Product Class: Other

α1-2,4,6 Fucosidase O
NEBU cloned at NEB recombinant 37 65 Heat

Product Introduction

α1-2,4,6 Fucosidase O is a broad specificity exoglycosidase that catalyzes the hydrolysis of terminal α1-2, α1-4 and α1-6 linked fucose residues from oligosaccharides. α1-2,4,6 Fucosidase O cleaves α1-6 fucose residues more efficiently than other linkages.

  • Recombinant enzyme with no detectable endoglycosidase or other exoglycosidase contaminating activities
  • Glycerol -free for optimal performance in HPLC and mass spectrometry analysis
  • ≥95% purity, as determined by SDS-PAGE and intact ESI-MS
  • Optimal activity and stability for up to 12 months
Catalog # Size Concentration
P0749S 80.0 units 2000 units/ml
P0749L 400.0 units 2000 units/ml

Product Information

Description

α1-2,4,6 Fucosidase O is a broad specificity exoglycosidase that catalyzes the hydrolysis of terminal α1-2, α1-4 and α1-6 linked fucose residues from oligosaccharides. α1-2,4,6 Fucosidase O cleaves α1-6 fucose residues more efficiently than other linkages.
• Recombinant enzyme with no detectable endoglycosidase or other exoglycosidase contaminating activities
• Glycerol-free foroptimal performance in HPLC and mass spectrometry analysis
• ≥95% purity, as determined by SDS-PAGE and intact ESI-MS
• Optimal activity and stability for up to 12 months

Product Source

Cloned from Omnitrophica bacterium and expressed in E. coli.

Specificity

P0749 Specificity
This product is related to the following categories:
Exoglycosidases Products,
Proteome Analysis Products,
This product can be used in the following applications:
Glycan Sequencing,
Proteomics,
Recombinant Glycoprotein Expression,
Glycoprotein Analysis

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • P0749S     4    
      α1-2,4,6 Fucosidase O P0749SVIAL 4 1 x 0.04 ml 2,000 units/ml
      GlycoBuffer 1 B1727SVIAL -20 1 x 1 ml 10 X
  • P0749L     4    
      α1-2,4,6 Fucosidase O P0749LVIAL 4 1 x 0.2 ml 2,000 units/ml
      GlycoBuffer 1 B1727SVIAL -20 1 x 1 ml 10 X

Properties & Usage

Unit Definition

One unit is defined as the amount of enzyme required to cleave > 95% of the fucose from 1 nmol of G0F from human IgG [GlcNAcβ1-2Manα1-6(GlcNAcβ1-2Manα1-3)Manβ1-4GlcNAcβ1-4GlcNAc(Fucα1-6)-AMAC], in 1 hour at 37°C in a total reaction volume of 10 µl.

Reaction Conditions

1X GlycoBuffer 1
Incubate at 37°C

1X GlycoBuffer 1
5 mM CaCl2
50 mM sodium acetate
(pH 5.5 @ 25°C)

Storage Buffer

20 mM Tris-HCl
50 mM NaCl
1 mM EDTA
pH 7.5 @ 20°C

Heat Inactivation

65°C for 10 minutes

Molecular Weight

Apparent: 49 kDa

Unit Assay Conditions

Two fold dilutions of α1-2,4,6 Fucosidase O are incubated with 1 nmol AMAC-labeled substrate and 1X GlycoBuffer 1 in a 10 µl reaction. The reaction mix is incubated at 37°C for 1 hour. Separation of reaction products are visualized via thin layer chromatography (1).

Product Notes

  1. The enzymatic unit definition is calculated using a 37°C reaction temperature, to accommodate for simultaneous glycosidase digestions and N-glycan sequencing protocols. However, the optimal reaction temperature for α1-2,4,6 Fucosidase O is 50°C. A 37°C reaction temperature results in ~35% lower activity.
  2. In order to achieve complete removal of core α1-6 fucose residues from glycans with instant labels, it may be necessary to use longer incubation times (18 hours).
  3. α1-2,4,6 Fucosidase O is only active on an intact antibody if it is trimmed to the trimannosyl core.
  4. Reactions may be scaled-up linearly to accommodate larger reaction volumes.

References

  1. Wong-Madden, S.T. and Landry, D. (1995). Glycobiology. 5, 19-28.

Protocols, Manuals & Usage

Protocols

  1. α1-2,4,6 Fucosidase O Digestion of Released Labeled Glycans Protocol

Application Notes

Tools & Resources

Web Tools

FAQs & Troubleshooting

FAQs

  1. What is the difference between α1-2,3,4,6 Fucosidase and α1-2,4,6 Fucosidase O?
  2. Can α1-2,4,6 Fucosidase O cleave fucose residues in the presence of instant labelled glycans?
  3. Can I use α1-2,4,6 Fucosidase O in a double digest with other exoglycosidases?
  4. Can α1-2,4,6 Fucosidase O cleave α1-3 fucose residues?
  5. Can α1-2,4,6 Fucosidase O cleave branched and complex substrates?
  6. Can α1-2,4,6 Fucosidase O cleave fucose residues from an intact antibody?
  7. What is the optimal incubation temperature for α1-2,4,6 Fucosidase O?
  8. What is a good method to re-purify a glycan or glycopeptide after exoglycosidase treatment?
  9. How much exoglycosidase should be used?

Tech Tips

Repeated freeze/thaw cycles may reduce activity. Recommended storage temperature is 4°C.

The optimal pH range is 4.5 – 5.5.

If using the enzyme alone (without other glycosidases), recommended incubation temperature is 50°C.

Longer incubation times (4 hours to overnight) may be needed for digestion of complex, oligosaccharides with multiple branches.

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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