Recombinant Human 15-PGDH/HPGD Protein, CF

R&D Systems | Catalog # 5660-DH

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Key Product Details

  • R&D Systems E. coli-derived Recombinant Human 15-PGDH/HPGD Protein (5660-DH)
  • 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

P15428

Structure / Form

Noncovalently-linked homodimer

Applications

Enzyme Activity
View all 15-PGDH/HPGD Proteins and Enzymes »

Product Specifications

Source

E. coli-derived human 15-PGDH/HPGD protein
Met1-Gln266, with a C-terminal 6-His tag

Purity

>95%, 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

Met1

Predicted Molecular Mass

30 kDa

SDS-PAGE

28 kDa, reducing conditions

Activity

Measure by the production of NADH during the oxidation of PGF2 alpha.
The specific activity is >1,500 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

5660-DH
Formulation Supplied as a 0.2 μm filtered solution in HEPES, NaCl, DTT and Glycerol.
Shipping The product is shipped with dry ice or equivalent. 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, -70 °C as supplied.
  • 3 months, -70 °C under sterile conditions after opening.

Background: 15-PGDH/HPGD

HPGD, or 15-hydroxyprostaglandin dehydrogenase, is a NAD+-linked dehydrogenase that oxidizes the hydroxyl group at position 15 of prostaglandins to a ketone, resulting in a loss of biological activity (1). HPGD is a major enzyme for the catabolism of prostaglandins. The enzyme is a member of the short-chain alcohol dehydrogenase family of enzymes (2). HPGD is a cytosolic enzyme expressed in most tissues, with highest expression levels in placenta, lung, and kidney (3). It is inhibited by aspirin and nonsteroidal anti-inflammatory drugs (4). Defects in HPGD are a cause of hypertrophic osteoarthropathy (5).

References

  1. Anggard, E. and B. Samuelsson (1964) J. Biol. Chem. 239:4097.
  2. Krook, M. et al. (1990) Biochemistry 29:738.
  3. Tai, H.H. (1976) Biochemistry 15:4586.
  4. Mak, O.T. et al. (1982) Biosci. Rep. 2:503.
  5. Uppal, S. et al. (2008) Nat. Genet. 40:789.

Long Name

15-Hydroxyprostaglandin Dehydrogenase

Alternate Names

15PGDH, HPGD, PGDH1, SDR36C1

Entrez Gene IDs

3248 (Human); 15446 (Mouse); 79242 (Rat)

Gene Symbol

HPGD

UniProt

P15428 (Human)

Additional 15-PGDH/HPGD Products

Product Documents for Recombinant Human 15-PGDH/HPGD Protein, CF

Certificate of Analysis

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Note: Certificate of Analysis not available for kit components.

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Product Specific Notices for Recombinant Human 15-PGDH/HPGD Protein, CF

For research use only

Citations for Recombinant Human 15-PGDH/HPGD Protein, CF

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Protocols

View specific protocols for Recombinant Human 15-PGDH/HPGD Protein, CF (5660-DH):

Materials
  • Assay Buffer: 50 mM Tris, 100 mM NaCl, 2 mM Dithiothreitol (DTT), pH 9.0
  • Recombinant Human 15‑PGDH/HPGD (rhHPGD) (Catalog # 5660-DH)
  • beta -Nicotinamide adenine dinucleotide ( beta -NAD) (Sigma, Catalog # N6522), 100 mM stock in deionized water
  • Prostaglandin F2 alpha (PGF2 alpha ) (Sigma, Catalog # P0424), 10 mM stock in absolute ethanol
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Prepare the Substrate Mixture.
    1. Dilute beta -NAD to 1 mM in Assay Buffer.
    2. Dilute PGF2 alpha to 0.4 mM in Assay Buffer.
    3. Mix equal volumes of each for a final concentration of 0.5 mM beta -NAD and 0.2 mM PGF2 alpha.
  2. Dilute rhHPGD to 1.0 ng/μL in Assay Buffer.
  3. Load in a plate 50 μL of 1.0 ng/μL rhHPGD, and start the reaction by adding 50 μL of Substrate Mixture.
  4. Include a Substrate Blank containing 50 μL of Assay Buffer and 50 μL of Substrate Mixture.
  5. Read at 339 nm in kinetic mode for 5 minutes.

     Specific Activity (pmol/min/µg) =

 Adjusted Vmax* (OD/min) x well volume (L) x 1012 pmol/mol
ext. coeff** (M-1cm-1) x path corr.*** (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank 
     **Using the extinction coefficient 6220 M-1cm-1 
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Well:
  • rhHPGD: 0.050 µg
  • beta -NAD: 0.25 mM
  • PGF2 alpha : 0.1 mM

FAQs for Recombinant Human 15-PGDH/HPGD Protein, CF

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  • Q: Why am I observing an additional higher molecular weight band?

    A: We expect this higher molecular weight band to be the homodimer. In this homodimer, the two monomers are noncovalently associated with each other. Since they are not joined by disulfide bonds, reducing conditions are not expected to separate the two monomers.

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