Recombinant S. agalactiae Hyaluronan Lyase Protein, CF

Catalog # Availability Size / Price Qty
5150-GH-010
Product Details
Citations (1)
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Recombinant S. agalactiae Hyaluronan Lyase Protein, CF Summary

Purity
>95%, by SDS-PAGE under reducing conditions and visualized by silver stain
Endotoxin Level
<1.0 EU per 1 μg of the protein by the LAL method.
Activity
Measured by its ability to cleave the native substrate Hyaluronan (Catalog # GLR004). The specific activity is >75,000 pmol/min/µg, as measured under the described conditions.
Source
E. coli-derived s. agalactiae Hyaluronan Lyase protein
Ser259-Ile1072, with an N-terminal Met and 6-His tag
Accession # NP_688206
Accession #
N-terminal Sequence
Analysis
Met
Predicted Molecular Mass
93 kDa
SDS-PAGE
86 kDa, reducing conditions

Product Datasheets

5150-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.

Assay Procedure

Materials
  • Assay Buffer: 500 mM Sodium Acetate, 50 mM CaCl2, pH 6.5
  • Recombinant Streptococcus agalactiae Hyaluronan Lyase (rHyluronan Lyase) (Catalog # 5150-GH)
  • Substrate: Hyaluronan (Catalog # GLR004)
  • 96 Well UV Transparent Plate (Costar, Catalog # 3635)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute Substrate to 5.0 mg/mL in Assay Buffer.
  2. Dilute rHyaluronan Lyase to 2.0 µg/mL in Assay Buffer.
  3. Load into a plate 50 µL of 2.0 µg/mL rHyaluronan Lyase, and start the reaction by adding 50 µL of 5.0 mg/mL Substrate. Include a Substrate Blank containing 50 µL of Assay Buffer and 50 µL of 5.0 mg/mL Substrate.
  4. Read plate at 232 nm in kinetic mode for 5 minutes.
  5. Calculate specific activity:

     Specific Activity (pmol/min/µg) =

Adjusted Vmax* (OD/min) x well volume (L) x 10^12 pmol/M
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank
      **Using the extinction coefficient 3800 M-1cm-1
      ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rHyluronan Lyase: 0.1 µg
  • Substrate: 2.5 mg/mL

Reconstitution Calculator

Reconstitution Calculator

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Background: Hyaluronan Lyase

The hyaluronan lyase from Streptococci agalactiae is highly specific towards hyaluronan (1). However, a thousand fold lower activity towards unsulfated chondroitin sulfate has also been observed (2). The enzyme activity is Ca2+ dependent (3). During digestion the enzyme moves processively along the hyaluronan chains continuously releasing disaccharide units as it travels (1). The recombinant enzyme can be used to digest hyaluronan and study chondroitin sulfate structure (2). Unlike hyaluronidases, hyaluronan lyase digestion of the substrate creates a double bond at the non-reducing terminus of the product; therefore, the digestion progress can be followed by UV spectrometry and the enzyme can be used for hyaluronan quantification. The recombinant enzyme can also be used for drug screening because the enzyme is a major pathogenic factor for Streptococci agalactiae that causes serious, often fatal, neonatal infections (4). The expressed protein corresponds to the mature form of the native protein (3).

References
  1. Lin, B. et al. (1994) J. Biol. Chem. 269:30113.
  2. Baker, J. et al. (1997) Biochem. J. 327:65.
  3. Akhtar, M. et al. (2006) J. Biol. Chem. 281:28336.
  4. Hynes, W.L. and Walton, S.L. (2000) FEMS Microbiol. Lett. 183:201.
Entrez Gene IDs
3685697 (S. pombe)
Alternate Names
Hyaluronan Lyase

Citation for Recombinant S. agalactiae Hyaluronan Lyase Protein, CF

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

1 Citation: Showing 1 - 1

  1. Imaging specific cellular glycan structures using glycosyltransferases via click chemistry.
    Authors: Wu Z, Person A, Anderson M, Burroughs B, Tatge T, Khatri K, Zou Y, Wang L, Geders T, Zaia J, Sackstein R
    Glycobiology, 2017;0(0):.
    Species: Human
    Sample Types: Whole Cells
    Applications: Click Chemistry

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