Glyko® Sialidase A™ [GK80040]

Price:
$ 250.00

Description

Releases α(2,3)-, α(2,6)-, α(2,8)-, and α(2,9)-linked N-acetylneuraminic acid from oligosaccharides and glycoproteins. Also capable of releasing N-glycolylneuraminic acid.

Source:
Recombinant gene from Arthrobacter ureafaciens, expressed in E. coli.

Specificity:
The enzyme releases α(2,3)-, α(2,6)-, α(2,8)-, and α(2,9)-linked N-acetylneuraminic acid from complex carbohydrates. The initial rate of hydrolysis of α(2,6) linkages is reported to be approximately twice that of α(2,3)-linked sialic acid however, in practice, this kinetic selectivity is of little consequence during extended incubations. Effective digestion of glycolipid substrates is facilitated by addition of a detergent, such as sodium taurodeoxycholate to the incubation.

Sialidase A is capable of releasing N-glycolylneuraminic acid (Neu5Gc, NGNA) in addition to N-acetylneuraminic acid (Neu5Ac, NANA) [1], although similarly to other sialidases [2] the activity is lower toward Neu5Gc than Neu5Ac.

The enzyme can also be used to remove sialic acid from gangliosides [3], glycosphingolipds (ceramide & oligosaccharide) with sialic acid.

May be used for exoglycosidase sequencing in conjunction with Sialidase S (GK80021), which is specific for α(2,3)-linked sialic acids.

Ships with:
WS0049 5x Reaction Buffer B [250 mM sodium phosphate pH 6.0]

Reaction Buffer:
5X concentrated buffer which when diluted gives 50 mM sodium phosphate pH 6.0.

Formulation:
20 mM Tris HCl pH 7.5, containing 25 mM NaCl.

pH optimum:
6.0

Unit Definition:
One unit is defined as the amount of enzyme required to catalyze the release of 1 μmole of p-nitrophenol from p-nitrophenyl-α-D-N-acetylneuraminic acid per minute at 37° C, pH 5.5.

Size: 1 U (200 µl)

Concentration: ≥ 5 U/ml

Product Code: GK80040

  1. Uchida Y, Tsukada Y, Sugimori T. Enzymatic properties of neuraminidases from Arthrobacter ureafaciens.  1979 Nov;86(5):1573-85.
  2. Schauer R. Chemistry, metabolism, and biological functions of sialic acids.  1982;40:131-234.
  3. Larsson EA, Olsson U, Whitmore CD, Martins R, Tettamanti G, Schnaar RL, Dovichi NJ, Palcic MM, Hindsgaul O. Synthesis of reference standards to enable single cell metabolomic studies of tetramethylrhodamine-labeled ganglioside GM1.  2007 Feb 26;342(3-4):482-9
Tech Info

GK80040 Technical Data Sheet  

 

FAQs

Q. Does GK80040 contain a purification tag, such as 6xHis?

A. No. Although GK80040 Sialidase A is expressed recombinantly, it does not contain a purification tag.

Q. I do not want to use the sodium phosphate reaction buffer supplied with GK80040. Can I use an acetate buffer? Is this good for digestion with multiple exoglycosidases?

A. We use an ammonium acetate buffer when we run multi-enzyme exoglycosidases digests that include GK80040 Sialidase A. We suggest a 10X reaction buffer of 500mM ammonium acetate pH 5.5 w/ 0.05% azide. The reaction buffer can also be used diluted to 20X (25mM ammonium acetate) and lower with purified glycans. This buffer works with most of our exoglycosidases in an overnight digestion: GK80040, GK80021, GKX-5014, GKX-5013, GKX-5023, GKX-5007, GKX-5010. Please Contact Us for more details.

If the glycans need to purified from the reaction prior to analysis by e.g. LC-MS, samples can be spun through Nanosep® 10K Omega spin filters to remove protein. For data using these methods, please see our poster, Characterizing Low-Abundance Glycans on Therapeutic Proteins.

Also, acetate buffers work better than phosphate for injecting onto LC (phosphate phase separates in acetonitrile), and acetate is volatile so samples can be dried down if necessary.

Product Citations
  1. Tharakaraman K, Raman R, Viswanathan K, Stebbins NW, Jayaraman A, Krishnan A, Sasisekharan V, Sasisekharan R.  Structural determinants for naturally evolving H5N1 hemagglutinin to switch its receptor specificity.  2013 Jun 20;153(7):1475-85.
  2. Cohen M, Elkabets M, Perlmutter M, Porgador A, Voronov E, Apte RN, Lichtenstein RG.  Sialylation of 3-methylcholanthrene-induced fibrosarcoma determines antitumor immune responses during immunoediting.   2010 Nov 15;185(10):5869-78.
  3. Stefanich EG, Ren S, Danilenko DM, Lim A, Song A, Iyer S, Fielder PJ.  Evidence for an asialoglycoprotein receptor on nonparenchymal cells for O-linked glycoproteins.   2008 Nov;327(2):308-15.
  4. Larsson EA, Olsson U, Whitmore CD, Martins R, Tettamanti G, Schnaar RL, Dovichi NJ, Palcic MM, Hindsgaul O. Synthesis of reference standards to enable single cell metabolomic studies of tetramethylrhodamine-labeled ganglioside GM1.  2007 Feb 26;342(3-4):482-9.
  5. Malakhov MP, Aschenbrenner LM, Smee DF, Wandersee MK, Sidwell RW, Gubareva LV, Mshin VP, Hayden FG, Kim DH, Ing A, Campbell ER, Yu M, Fang F.  Sialidase fusion protein as a novel broad-spectrum inhibitor of influenza virus infection.   2006 Apr;50(4):1470-9.
  6. Zhang L, Bukreyev A, Thompson CI, Watson B, Peeples ME, Collins PL, Pickles RJ.  Infection of ciliated cells by human parainfluenza virus type 3 in an in vitro model of human airway epithelium.   2005 Jan;79(2):1113-24.
Safety

Product Safety Documentation for GK80040:

Product/Part No. Description
GK80040 Glyko® Sialidase A™ 
WS0049 5x Reaction Buffer B 

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