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Exoglycosidases >
α-N-Acetyl-galactosaminidase |
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Prices are in US dollars and valid only for US orders.
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Substrate Specificity:
Description:
α-N-Acetyl-galactosaminidase is a highly specific exoglycosidase that catalyzes the hydrolysis of α-linked D-N-acetyl-galactosamine residues from oligosaccharides and proteins (1).
Source:
Cloned from a proprietary strain and expressed in E. coli at NEB (1).
Reagents Supplied:
G7 Reaction Buffer (10X)
BSA (100X)
Enzyme Properties
Molecular Weight:
Apparent: 47 kDa
Reaction & Storage Conditions
Reaction Conditions:
1X G7 Reaction Buffer
Supplemented with 100 μg/ml Bovine Serum Albumin
Incubate at
37°C.
1X G7 Reaction Buffer:
50 mM sodium phosphate
pH 7.5 @ 25°C
Unit Definition:
One unit is defined as the amount of enzyme required to cleave > 95% of the terminal α-D-N-acetyl-galactosamine from 1 nmol (GalNAcα1-3)(Fucα1-2)Galα1-4Glc-7-amino-4-methyl-coumarin (AMC), in 1 hour at 37°C in a total reaction volume of 10 µl.
Two fold dilutions of α-N-Acetyl-galactosamininidase are incubated with 1 nmol AMC-labeled substrate in 1X G1 Buffer, supplemented with 100 µg/ml BSA, 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 (2).
Concentration:
20,000 units/ml
Storage Temperature:
-20°C
Avoid repeated freeze/thaw cycles.
Notes
Usage notes:- Optimal incubation times and enzyme concentrations must be determined empirically for a particular substrate.
FAQs
- How much exoglycosidase should be used?
- What is a good positive control for α-N-Acetyl-galactosaminidase?
- Do detergents inhibit exoglycosidases/endoglycosidases?
- Do detergents inhibit exoglycosidases/endoglycosidases?
- What are Glycosidases and their uses?
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Quality Assurance Statement:
No contaminating exoglycosidase or proteolytic activity could be detected.
Protease Activity:
After incubation of 300 units of α-N-Acetyl-galactosaminidase with a standard mixture of proteins
for 2 hours at 30ºC, no proteolytic activity could be detected by SDS-PAGE.
Glycosidase Assay:
20 units of α-N-Acetyl-galactosaminidase were incubated with 0.1 mM of flourescently-labeled oligosaccharides and glycopeptides, in a 10 µl reaction for 20 hours at 37°C (see list below). The reaction products were analyzed by TLC for digestion of substrate (3).
No other glycosidase activities were detected (ND) with the following substrates
β-N-Acetyl-glucosaminidase: GlcNAcβ1-4GlcNAcβ1-4GlcNAc-AMC ND
α-Fucosidase: Fucα1-2Galβ1-4Glc-AMC Galβ1-4 (Fucα1-3)GlcNAcβ1-3Galβ1-4Glc-AMC ND
β-Galactosidase: Galβ1-3GlcNAcβ1-4Galβ1-4Glc-AMC ND
α-Galactosidase: Galα1-3Galβ1-4Gal-AMC ND
α-Neuraminidase: Neu5Acα2-3Galβ1-3GlcNAcβ1-3Galβ 1-4Glc-AMC ND
α-Mannosidase: Manα1-6Manα1-6(Manα1-3)Man-AMC ND
β-Glucosidase: Glcβ1-4Glcβ1-4Glc-AMC ND
β-Xylosidase: Xylβ1-4Xylβ1-4Xylβ1-4Xyl-AMC ND
β-Mannosidase: Manβ1-4Manβ1-4Man-AMC ND
Endo F1, F2, H: Dansylated invertase high mannose. ND
Endo F2, F3: Dansylated fibrinogen biantennary. ND
PNGase F: Fluoresceinated fetuin triantennary. ND
References
- Landry, D. and Guthrie, E.P., New England Biolabs, unpublished observations.
- Wong-Madden, S.T. and Landry, D. (1995) Glycobiology, 5, 19-28.
Reagents Sold Separately
BSA
Companion Products
Exoglycosidase Reaction Buffer Pack
Legal
Patents:
New England Biolabs, Inc. : U.S. Patent No. 6,423,525
New England Biolabs, Inc. : U.S. Patent No. 6,458,573
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