Product Class: Other

β-N-Acetylhexosaminidasef
NEBU cloned at NEB recombinant 37 75 Heat

New Reaction Buffer

Product Introduction

β-N-Acetylhexosaminidasef is an exoglycosidase that catalyzes the hydrolysis of terminal, non-reducing β-N-Acetylgalactosamine and glucosamine residues from oligosaccharides.

  • Can be used for Epigenetic applications to remove O-GlcNAc from serine/threonine residues on transcription factors, histones, RNA-binding proteins and other O-GlcNAc modified proteins
  • 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 24 months
Catalog # Size Concentration
P0721S 500.0 units 5000 units/ml
P0721L 2500.0 units 5000 units/ml

Product Information

Description

Highlights

  • Epigenetics applications:  O-GlcNac (N-Acetylhexosamine attached to serine or threonine) is a regulator of nuclear protein function. β-N-Acetylhexosaminidasef removes O-GlcNAc from transcription factors, histones, RNA-binding proteins, etc.
β-N-Acetylhexosaminidasef is a recombinant protein fusion of β-N-Acetylhexosaminidase (1) and maltose binding protein. It has identical activity to β-N-Acetylhexosaminidase. β-N-Acetylhexosaminidasef catalyzes the hydrolysis of terminal β-D-N-acetyl-galactosamine and glucosamine residues from oligosaccharides.  β-N-Acetylhexosaminidasef can be used to remove O-GlcNAc (N-Acetylglucosaminidase on serine/threonine)(4). 

Substrate Specificity:

Product Source

Cloned from Streptomyces plicatus and overexpressed in E. coli.
This product is related to the following categories:
Exoglycosidases Products
This product can be used in the following applications:
Glycan Sequencing,
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
  • P0721S     -20    
      β-N-Acetylhexosaminidasef P0721SVIAL -20 1 x 0.1 ml 5,000 units/ml
      GlycoBuffer 1 B1727SVIAL -20 1 x 1 ml 10 X
  • P0721L     -20    
      β-N-Acetylhexosaminidasef P0721LVIAL -20 1 x 0.5 ml 5,000 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 terminal β-D-N-Acetylgalactosamine from 1 nmol of GalNAcβ1-4Galβ1-4Glc-7-amino-4-methyl-coumarin (AMC), 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

50 mM NaCl
20 mM Tris-HCl
5 mM EDTA
pH 7.5 @ 25°C

Heat Inactivation

75°C for 10 minutes

Molecular Weight

Theoretical: 100000 daltons

Terminal Monosacaride

  • N-Acetyl-galactosamine and N-Acetyl-glucosamine Residues

Unit Assay Conditions

Two fold dilutions of β-N-Acetyl-hexosaminidasef are incubated with 1 nmol AMC-labeled substrate in 1X GlycoBuffer 1 in a 10 µl reaction. The reaction mix is incubated for 1 hour at 37°C. Separation of reaction products are visualized via thin layer chromatography (3).

References

  1. Robbins, P. et al. (1992). Gene. 111, 69-76.
  2. Guan, C. and Wong, S. New England Biolabs.. Unpublished observation
  3. Wong-Madden, S.T. and Landry D. (1995). Glycobiology. 5, 19-28.
  4. Magnelli, P.E. et al Methods Molecular Biology. 801, 189-211.

Protocols, Manuals & Usage

Protocols

  1. Typical Reaction Conditions for β-N-Acetylhexosaminidasef (P0721)

Usage & Guidelines

Application Notes

FAQs & Troubleshooting

FAQs

  1. How much exoglycosidase should be used?
  2. Do detergents inhibit exoglycosidases/endoglycosidases?
  3. What is a good positive control for β-N-Acetylhexosaminidasef?
  4. What are Glycosidases and their uses?
  5. Why have the NEB Glycosidase enzymes changed reaction buffers? What are the new reaction buffers and can I still use an enzyme with its old buffer? Where can I find the composition of the old buffers?
  6. How much exoglycosidase should be used?

Tech Tips

A good positive control for ßHexf is pNP-GlcNAc.

Using 1-2 µl is a good starting point for a 1 hr incubation of 1µg of glycoprotein or 100 nM of oligosaccharide.

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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