Endoglycosidase H is a recombinant glycosidase which cleaves within the chitobiose core of high mannose and some hybrid oligosaccharides from N-linked glycoproteins. A tagged version, which is a recombinant protein fusion of Endoglycosidase H and maltose binding protein is also available, Endo Hf (NEB #P0703).
Recombinant enzyme with no detectable exoglycosidase or other endoglycosidase 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
Product Information
Endoglycosidase H is a recombinant glycosidase which cleaves within the chitobiose core of high mannose and some hybrid oligosaccharides from N-linked glycoproteins.
60 µg of RNase B was incubated with 3,000 units of Endo H or Endo Hf under standard assay conditions
Aliquots were removed at various time points and measured for released carbohydrate. [Dubois et al. (1956) Anal. Chem. 28, 350-356].Mobility Shift Analysis 1 unit of Endo H, Endo Hf or PNGase F was incubated per 10 µg of RNase B under standard assay conditions. At various time points, aliquots were removed and analyzed on a 10-20% SDS-PAGE gel for carbohydrate (CHO) release. 1 unit is defined as the amount of enzyme required to remove >95% of the carbohydrate from 10 µg of RNase B in one hour at 37°C.
Product Source
Cloned from Streptomyces plicatus (2) and overexpressed in E.coli (3).
This product is related to the following categories:
The following reagents are supplied with this product:
NEB #
Component Name
Component #
Stored at (°C)
Amount
Concentration
P0702S
-20
Endo H
P0702SVIAL
-20
1 x 0.02 ml
500,000 units/ml
GlycoBuffer 3
B1720SVIAL
-20
1 x 1 ml
10 X
Glycoprotein Denaturing Buffer
B1704SVIAL
-20
1 x 1 ml
10 X
P0702L
-20
Endo H
P0702LVIAL
-20
1 x 0.1 ml
500,000 units/ml
GlycoBuffer 3
B1720SVIAL
-20
1 x 1 ml
10 X
Glycoprotein Denaturing Buffer
B1704SVIAL
-20
1 x 1 ml
10 X
Properties & Usage
Unit Definition
One unit is defined as the amount of enzyme required to remove > 95% of the carbohydrate from 10 µg of denatured RNase B in 1 hour at 37°C in a total reaction volume of 10 µl (10 NEB units = 1 IUB milliunit).
Unit Definition Assay: 10 µg of RNase B are denatured with 1X Glycoprotein Denaturing Buffer at 100°C for 10 minutes. After the addition of 1X GlycoBuffer 3, two-fold dilutions of Endo H are added and the reaction mix is incubated for 1 hour at 37°C. Separation of reaction products is visualized by SDS-PAGE.
1X Glycoprotein Denaturing Buffer 0.5% SDS 40 mM DTT
To deglycosylate a native glycoprotein, longer incubation time as well as more enzyme may be required.
Activity at different temperatures (measured after a 1 hour incubation of glycosidase and denatured RNase B at the given temperature): 37°C - 100%; 30°C - 65%; 25°C - 40%; 17°C - 25% and 2°C - 0%.
Typical reaction conditions: Please see FAQs.
References
Maley, F. et al. (1989). Anal. Biochem. 180, 195-204.
Robbins, P. et al. (1984). J. Biol. Chem. 259, 7577-7583.
Guan, C. and Wong,S. New England Biolabs, unpublished observations.
Arakel EC, Brandenburg S, Uchida K, Zhang H, Lin YW, Kohl T, Schrul B, Sulkin MS, Efimov IR, Nichols CG, Lehnart SE, Schwappach B (2014) Tuning the electrical properties of the heart by differential trafficking of KATP ion channel complexes J Cell Sci; 127(Pt 9), 2106-19. PubMedID: 24569881, DOI: 10.1242/jcs.141440
Möykkynen T, Coleman SK, Semenov A, Keinänen K (2014) The N-terminal domain modulates α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor desensitization J Biol Chem; 289(19), 13197-205. PubMedID: 24652293, DOI: 10.1074/jbc.M113.526301
Rosenbaum EE, Vasiljevic E, Brehm KS, Colley NJ (2014) Mutations in four glycosyl hydrolases reveal a highly coordinated pathway for rhodopsin biosynthesis and N-glycan trimming in Drosophila melanogaster PLoS Genet; 10(5), e1004349. PubMedID: 24785692, DOI: 10.1371/journal.pgen.1004349
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Specifications & Change Notifications
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