Recombinant Human HO-2/HMOX2 Protein, CF

R&D Systems | Catalog # 3170-HM

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Key Product Details

  • R&D Systems E. coli-derived Recombinant Human HO-2/HMOX2 Protein (3170-HM)
  • 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

P30519

Applications

Enzyme Activity
View all HO-2/HMOX2 Proteins and Enzymes »

Product Specifications

Source

E. coli-derived human HO-2/HMOX2 protein
Ser2-Leu291, with a C-terminal 6-His tag

Purity

>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane.

Endotoxin Level

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

N-terminal Sequence Analysis

Ser2

Predicted Molecular Mass

34 kDa

SDS-PAGE

31 & 35 kDa, reducing conditions

Activity

Measured by its ability to oxidize hemin to biliverdin.
The specific activity is >3.5 pmol/min/μg, as measured under the described conditions.

Formulation, Preparation, and Storage

3170-HM
Formulation Supplied as a 0.2 μm filtered solution in Tris and NaCl.
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: HO-2/HMOX2

Heme oxygenase (HO) is the rate limiting enzyme in heme catabolism (1). It cleaves heme to biliverdin, carbon monoxide, and iron; biliverdin is subsequently converted by biliverdin reductase to bilirubin. The mechanism of HO is unique in that heme serves as both the substrate of the enzyme and as the prosthetic group for the activation of iron-bound O2. HO activity is highest in spleen where senescent erythrocytes are sequestered and destroyed. Two isoforms, HO-1 and HO-2, are expressed in most tissues. HO-1 is an inducible enzyme in response to heavy metals, oxidative stress, cytokines, and many drugs (2). HO-2 is constitutively expressed in the brain and testes. HO-1 is expressed mainly in spleen, liver, and kidney. In addition to its activity in the heme degradation pathway, HO-2 potentially serves as an oxygen sensor through its heme regulating motifs, which have the core sequence of Cys-Pro (3).

References

  1. Nader, G. (2008) Pharmacol. Rev. 60:79.
  2. Otterbein, L.E. and A.M.K. Choi (2000) Am. J. Physiol. Lung Cell Mol. Physiol. 279:1029.
  3. Yi. L. et al. (2009) J. Biol. Chem. 284:20556.

Long Name

Heme Oxygenase 2

Alternate Names

HMOX2, HO2

Entrez Gene IDs

3163 (Human); 15369 (Mouse); 79239 (Rat)

Gene Symbol

HMOX2

UniProt

P30519

Additional HO-2/HMOX2 Products

Product Documents for Recombinant Human HO-2/HMOX2 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 HO-2/HMOX2 Protein, CF

For research use only

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Protocols

View specific protocols for Recombinant Human HO-2/HMOX2 Protein, CF (3170-HM):

Materials
  • Assay Buffer: 50 mM Tris, pH 7.5
  • Recombinant Human HO‑2/HMOX2 (rhHO‑2) (Catalog # 3170-HM)
  • Recombinant Human POR/Cytochrome P450 Reductase (rhPOR) (Catalog # 6340-PR)
  • Recombinant Human Biliverdin Reductase A/BLVRA (rhBLVRA) (Catalog # 6454-BR)
  • Catalase (Sigma, Catalog # C30), 100,000 U/mL stock
  • Hemin (Sigma, Catalog # H9039), 10 mM stock in DMSO
  • Bovine Serum Albumin (BSA), 100 mg/mL stock in deionized water
  • beta -Nicotinamide adenine 2'-phosphate reduced tetrasodium salt hydrate (NADPH) (Sigma, Catalog # N7505), 20 mM stock in deionized water
  • 96-well Clear Plate (Costar, Catalog # 92592)
  • Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent
  1. Dilute Hemin to 1 mM in Assay Buffer.
  2. Prepare Reaction Mixture containing 60 µM Hemin, 4 mg/mL BSA, 1000 U/mL catalase, 80 µg/mL rhPOR, and 80 µg/mL rhBLVRA in Assay Buffer.
  3. Prepare 80 µg/mL rhHO-2 in Assay Buffer.
  4. Dilute NADPH to 1 mM in Assay Buffer.
  5. In a clear plate, load 25 µL dilute rhHO-2 and add 25 µL Reaction Mixture. Include an enzyme blank containing 25 µL Assay Buffer and 25 µL Reaction Mixture.
  6. Start the reaction by adding 50 µL 1 mM NADPH to all wells used.
  7. Read absorbance at 468 nm (bottom read) in kinetic mode for 5 minutes.
  8. Calculate specific activity:

     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 43500 M-1cm-1 
     ***Using the path correction 0.32 cm
     Note: the output of many spectrophotometers is in mOD Per Reaction:

  • rhHO-2: 2 µg
  • rhPOR: 2 µg
  • rhBLVRA: 2 µg
  • Catalase: 250 U/mL
  • Hemin: 15 µM
  • BSA: 1 mg/mL
  • NADPH: 0.5 mM

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