Recombinant S. pyogenes Endo S2 His-tag Protein, CF

R&D Systems | Catalog # 10976-GH

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Key Product Details

  • R&D Systems E. coli-derived Recombinant S. pyogenes Endo S2 His-tag Protein (10976-GH)
  • Quality control testing to verify active proteins with lot specific assays by in-house scientists
  • All R&D Systems proteins are covered with a 100% guarantee

Source

E. coli

Accession Number

ACI61688.1

Applications

Enzyme Activity
View all Endo-beta-N-acetylglucosaminidase S2/Endo S2 Proteins and Enzymes »

Product Specifications

Source

E. coli-derived s. pyogenes Endo-beta-N-acetylglucosaminidase S2/Endo S2 protein
Glu37-Asp843, with a N-terminal Met & 6-His tag

Purity

>90%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

N-terminal Sequence Analysis

Met

Predicted Molecular Mass

92 kDa

SDS-PAGE

86-93 kDa, under reducing conditions.

Activity

Measured by its ability to digest Cy5-Labeled Glycan G2
>50% of Cy5-Labeled Glycan G2 (0.2 pmol) is digested by 0.2 μg of rSp. Endo-S2, as measured under the described conditions.

Scientific Data Images for Recombinant S. pyogenes Endo S2 His-tag Protein, CF

Recombinant Sp. Endo S2 (Endo S2) Enzyme Activity Diagram

Endo S2 recognizes the conserved N-glycans on the Fc region of IgG by hydrolyzing the chitobiose core (at the  beta -1,4 linkage between the two N-acetylglucosamines) of the N-glycans. Endo S2 leaves one GlcNAc residue attached the the asparagine of the peptide backbone. R1 and R2 can be oligosaccharide extensions containing Gal, GlcNAc, and Sialic Acid. R3 can be unmodified or Core-6 Fucose.

Recombinant S. pyogenes Endo S2 SDS-PAGE

1 μg/lane of rSp. Endo-S2 (Catalog # 10976-GH) was resolved with SDS-PAGE under reducing (R) conditions and visualized by silver staining, showing a band at 92 kDa.

Fluorescent Gel Mobility Shift caused by rSp. Endo S2.

Lane 1 contained substrate Cy5-labeled G2 GL302. In the presence of Endo S2, the glycan was digested to products and the smaller product Fuc-alpha,6-GlcNAc is observed.

Formulation, Preparation, and Storage

10976-GH
Formulation Supplied as a 0.2 μm filtered solution in Tris and NaCl.
Shipping The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.

Background: Endo-beta-N-acetylglucosaminidase S2/Endo S2

Streptococcus pyogenes is a leading Gram-positive bacterial pathogen that can abolish the effector functions of human immunoglobulin G (IgG) through deglycosylation (1). Upon infection, the pathogen secret two endoglycosidases, Endo S and Endo S2, that specifically deglycosylate the conserved N-glycans on the Fc region of IgG by hydrolyzing the chitobiose core (at the beta -1,4 linkage between the two N-acetylglucosamines) of the N-glycans (2, 3). Endo S and S2 cleavage leave one GlcNAc residue remaining attached to the asparagine residue on the peptide backbone. The enzymes are highly specifc to native IgG molecules (3), suggesting that the local conformation of IgG is required for the enzymatic recognition. In comparison, PNGase F from Flavobacterium meningosepticum completely removes glycans from glycoproteins and is more active on denatured glycoproteins. Cleavage of the glycans from IgG antibodies by Endo S and S2 result in conformation change of the antibodies thereby dramatically diminish the binding affinity to their receptors (4) and abolish their opsonizing functions (5, 6). By using fluorophore labeled N-Glycans as substrates, we found that Endo S2 has similar activity to Endo S towards non-galactosylated IgG glycan species including oligomannose and hybrid glycans but shows much high activity on galactosylated and sialylated IgG glycan species. We also found that both enzymes are highly active on core-6 fucosylated IgG glycans but with no activities on those with bisecting GlcNAc.

References

  1. Nizet, V. (2007) J. Allergy Clin. Immunol. 120:13.
  2. Collin, M. and Olsen, A. (2001) EMBO J. 20:3046.
  3. Sjogren, J. et al. (2013) Biochem. J. 455:107.
  4. Allhorn, M. et al. (2008) PLoS ONE. 3:e1413.
  5. Collin, M. et al. (2002) Infect Immun. 70:6646.
  6. Sjögren, J. et al. (2011) BMC Microbiol. 11:120.

Long Name

Endo-beta-N-acetylglucosaminidase S2

Alternate Names

EndoS2, NdoS2

Gene Symbol

NdoS2

UniProt

ACI61688.1

Additional Endo-beta-N-acetylglucosaminidase S2/Endo S2 Products

Product Documents for Recombinant S. pyogenes Endo S2 His-tag Protein, CF

Certificate of Analysis

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Product Specific Notices for Recombinant S. pyogenes Endo S2 His-tag Protein, CF

For research use only

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Protocols

View specific protocols for Recombinant S. pyogenes Endo S2 His-tag Protein, CF (10976-GH):

Materials
  • Assay Buffer: 50 mM MES, pH 6.0
  • Recombinant Sp. Endo-S2 (rSpEndo-S2) (Catalog # 10976-GH)
  • Cy5-Labeled Glycan G2 (Cy5-G2) (Catalog # GL302)
  • 15% SDS-PAGE gel and SDS-PAGE Reagents
  • Reducing SDS-PAGE Sample Buffer
  • Fluorescent imager
  1. Dilute rSpEndo-S2 to 20 ng/µL in Assay Buffer.
  2. Dilute Cy5-G2 to 0.02 µM in Assay Buffer.
  3. Prepare reaction by combine 10 µL of diluted rSpEndo-S2 and 10 µL of diluted Cy5-G2.
  4. Prepare a negative control by combining 10 µL of diluted Cy5-G2 and 10 µL of Assay Buffer.
  5. Incubate reaction(s) and negative control at 37 °C for 60 minutes.
  6. Add 7 µL of reducing SDS-PAGE sample buffer to each reaction and negative control.
  7. Load 13.5 µL of each sample and control per well on a 15% SDS-PAGE gel and run SDS-PAGE until the dye front has migrated more than two thirds of the way down the gel.
  8. Acquire gel image with a fluorescent imager.
  9. Analyze the amount (%) of Cy5-G2 digested by rSpEndo-S2 in each reaction lane.

Per Reaction:

  • rSpEndo-S2:  0.20 µg
  • Cy5-G2: 0.2 pmol





















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