N-Glycanase® (PNGase F), 200 mU [GKE-5006B]

Price:
$ 446.00

Description

PNGase F, peptide-N-glycosidase F, peptide-N4-(N-acetyl-β-glucosaminyl)asparagine amidase. Releases intact N-glycans by cleaving between the innermost GlcNAc and Asn. 

Source:
Recombinant gene from Elizabethkingia meningosepticum, expressed in E. coli. The source organism was previously known as Chryseobacterium [Flavobacterium] meningosepticum [1].

Specificity: 
Cleaves all N-linked complex, hybrid or high mannose oligosaccharides, unless α(1-3) core fucosylated, as in plant glycans and some insect glycans. Asparagine must be peptide-bonded at both termini. Phosphate, sulfate and sialic acid groups attached to the oligosaccharide do not affect cleavage. Endo F free. 

 

Figure 1: Cleavage specificity for GKE-5006 N-Glycanase (PNGase F).
R1: N & C substitution other than H
R2: may be H or the rest of an oligosaccharide
R3: may be H or α(1-6)-linked fucose

Applications:

  • Release of intact N-linked glycans from glycopeptides and glycoproteins
  • Structure-function studies of N-glycosylated glycoproteins
  • Preparation of deglycosylated proteins for molecular weight estimation or crystallography studies

Supplied at a concentration of ≥ 2.5 U/ml. For a more concentrated formulation of ≥ 10 U/ml, which may be useful for native digestions, see GKE-5010B or GKE-5020B (EDTA-free).

    "A & B" pack sizes ship with:

      • 1 ea WS0010 5x N-Glycanase Reaction Buffer
      • 1 ea WS0012 Denaturation Solution
      • 1 ea WS0013 Detergent Solution
      • 1 ea WS0145 5x N-Glycanase Tris Reaction Buffer (MS applications)

    "D" pack size does not ship with the above buffers.

      Formulation:
      20 mM Tris-HCl, 50 mM NaCl, 1 mM EDTA (pH 7.5)

      Reaction Buffer:
      5X concentrated buffer, when diluted gives 20 mM sodium phosphate pH 7.5, containing 0.02% sodium azide.

      MW:
      ~35,000 daltons

      pH optimum/range:
      8.6 / 7.5 - 9.5

      Unit Definition:
      One unit is defined as the amount of enzyme required to catalyze the release of N-linked oligosaccharides from 1 μmole of denatured ribonuclease B per minute at 37°C, pH 7.5.

      Not formulated for FACE® analysis, use GK50050 instead.

      Size: 200 mU in 80 µl

      Concentration: ≥ 2.5 U/ml

      Product Code: GKE-5006B

      1. Kim KK, Kim MK, Lim JH, Park HY, Lee ST. Transfer of Chryseobacterium meningosepticum and Chryseobacterium miricola to Elizabethkingia gen. nov. as Elizabethkingia meningoseptica comb. nov. and Elizabethkingia miricola comb. nov.  2005 May;55(Pt 3):1287-93.

      Tech Info

      GKE-5006B Technical Data Sheet  

      FAQs

      Q. You have a number of PNGase F products. What are the differences?

      A. We carry a number of different formulations and pack sizes of PNGase F:

      Product Pack size Volume Concentration* Recombinant Native EDTA
      GKE-5006 A - 100 mU 40 µl ≥ 2.5 U/ml 1mM
      B - 200 mU 80 µl
      D - 1 U† 400 µl
      GKE-5016 A - 100 mU 40 µl ≥ 2.5 U/ml 0
      B - 200 mU 80 µl
      D - 1 U 400 µl
      GKE-5010 (PLUS) B - 400 mU 40 µl ≥ 10 U/ml 1 mM
      D - 1 U 100 µl
      GKE-5020 (ULTRA) B - 400 mU 40 µl ≥ 10 U/ml 0
      D - 1 U 100 µl
      GKE-5003 100 mU 50 µl ≥ 2.0 U/ml 5 mM

      *One unit of N-Glycanase is defined as the amount of enzyme required to catalyze the release of N-linked oligosaccharides from 1 μmole of denatured ribonuclease B per minute at pH 7.5 and 37°C.

      †Buffers and denaturant are not supplied with 1 U "D" pack sizes, but can be made available on request.

      EDTA was included historically in native PNGase F formulations as a stabilizer. For the GKE-5020 formulation we revisited the idea, due to requests from customers for a PNGase F formulation without EDTA. We believe that stability is not as much of a concern with the recombinant form of the enzyme.

      Q. I am running MS of proteins with and without PNGase F treatment. Is there a +1 Da change when using PNGaseF in addition to the MW loss of the N-linked glycan?

      A. Yes, a + 1 Da change on the protein is to be expected when deglycosyalting with PNGase F. The process of PNGase F removing the N-glycans involves the deamination of asparagine to aspartic acid. We have some references on this in our tech data sheet for PNGase F.

      Asparagine (N): 132.12 Da

      Aspartic acid (D): 133.10 Da

      Safety

      Product Safety Documentation for GKE-5006B:

      Product/Part No. Description
      GKE-5006B

      US-N-Glycanase®

      GB-N-Glycanase®

      

      WS0010 5x N-Glycanase Reaction Buffer
      WS0012 Denaturation Solution
      WS0013 Detergent Solution
      WS0145 5x N-Glycanase Tris Reaction Buffer

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