P8415
Peroxidase from horseradish
Type XII, essentially salt-free, lyophilized powder, ≥250 units/mg solid (using pyrogallol)
Synonym(s):
Donor:hydrogen-peroxide oxidoreductase, Horseradish peroxidase
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About This Item
CAS Number:
NACRES:
NA.54
UNSPSC Code:
12352204
EC Number:
232-668-6
MDL number:
eCl@ss:
32160410
Specific activity:
≥250 units/mg solid (using pyrogallol)
Pricing and availability is not currently available.
type
Type XII
Quality Segment
form
essentially salt-free, lyophilized powder
specific activity
≥250 units/mg solid (using pyrogallol)
mol wt
~44 kDa
solubility
0.1 M phosphate buffer: soluble (pH 6.0), H2O: soluble
absorbance ratio
RZ ≥3.0
storage temp.
2-8°C
SMILES string
[O+H2]O[O-]
InChI
1S/H2O3/c1-3-2/h1-2H
InChI key
JSPLKZUTYZBBKA-UHFFFAOYSA-N
General description
HRP is a single chain polypeptide containing four disulfide bridges. It is a glycoprotein containing 18% carbohydrate. The carbohydrate composition consists of galactose, arabinose, xylose, fucose, mannose, mannosamine, and galactosamine depending upon the specific isozyme. Its molecular weight (~44 kDa) includes the polypeptide chain (33,890 Daltons), hemin plus Ca2+ (~700 Daltons), and carbohydrate (~9,400 Daltons). At least seven isozymes of HRP exist. The isoelectric point for horseradish peroxidase isozymes ranges from 3.0 - 9.0.
Horseradish peroxidase is isolated from horseradish roots (Amoracia rusticana) and belongs to the ferroprotoporphyrin group of peroxidases. HRP is a single chain polypeptide containing four disulfide bridges. It is a glycoprotein containing 18% carbohydrate. The carbohydrate composition consists of galactose, arabinose, xylose, fucose, mannose, mannosamine, and galactosamine depending upon the specific isozyme. Its molecular weight (~44 kDa) includes the polypeptide chain (33,890 Daltons), hemin plus Ca2+ (~700 Daltons), and carbohydrate (~9,400 Daltons). At least seven isozymes of HRP exist. The isoelectric point for horseradish peroxidase isozymes ranges from 3.0 - 9.0.
Application
Peroxidase from horseradish has been used to initiate peroxidase-coupled assay. It has also been used in the preparation of β-galactosidase (β-gal) stock solution.
Horseradish peroxidase (HRP) is isolated from horseradish roots (Amoracia rusticana) and belongs to the ferroprotoporphyrin group of peroxidases. It is used in biochemistry applications such as western blots, ELISA and Immunohistochemistry. Horseradish peroxidase is used to amplify a weak signal and increase detectability of a target molecule, such as a protein. Product P8415, type XII, is an essentially salt free lyophilized powder. It is a further purification of product P8375. It is commonly used to determine amounts of glucose and peroxides in solution. It has been used in an aspergillus fumigatus rapid susceptibility assay[1].
The enzyme has been used to determine H2O2 production in tobacco BY-2 cells using a spectrofluorimetric method.[2]
Biochem/physiol Actions
HRP readily combines with hydrogen peroxide (H2O2) and the resultant [HRP-H2O2] complex can oxidize a wide variety of hydrogen donors. The optimal pH is 6.0-6.5 and the enzyme is most stable in the pH range of 5.0-9.0. HRP can be conjugated to antibodies by several different methods including glutaraldehyde, periodate oxidation, through disulfide bonds, and also via amino and thiol directed cross-linkers. It is smaller and more stable than the enzyme labels β-galactosidase and alkaline phosphatase and hence, it is the most desired label. Also, its glycosylation leads to lower non-specific binding. It is also used for the determination of glucose and peroxides in solution.
When incubated with a substrate, horseradish peroxidase produces a coloured, fluorimetric, or luminescent derivative of the labeled molecule, allowing quantification. Horseradish peroxidase has been shown to slightly reduce the level of inhibition in a cydAB mutant. Known inhibitors are sodium azide, cyanide, L-cystine, dichromate, ethylenethiourea, hydroxylamine, sulfide, vanadate, p-aminobenzoic acid, and Cd2+, Co2+, Cu2+, Fe3+, Mn2+, Ni2+, and Pb2+ ions.
Preparation Note
Chromatographically purified
Analysis Note
Preliminary isoelectric focusing data indicates this is primarily isoenzyme C
The RZ (Reinheitszahl) is the absorbance ratio A403/A275 determined at 0.5-1.0 mg/ml in deionized water. It is a measure of hemin content, not enzymatic activity. Even preparations with high RZ may have low enzymatic activity.
Other Notes
A further purification of Peroxidase TypeVI (P8375).
One pyrogallol unit will form 1.0 mg purpurogallin from pyrogallol in 20 sec at pH 6.0 at 20 °C.
View more information on peroxidase at www.sigma-aldrich.com/enzymeexplorer.
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This Item | |||
|---|---|---|---|
| specific activity ≥250 units/mg solid (using pyrogallol) | specific activity ~150 U/mg | specific activity ≥225 units/mg protein (biuret, using pyrogallol) | specific activity ≥200 U/mg |
| form essentially salt-free, lyophilized powder | form powder | form ammonium sulfate suspension | form lyophilized solid |
| storage temp. 2-8°C | storage temp. 2-8°C | storage temp. 2-8°C | storage temp. −20°C |
| mol wt ~44 kDa | mol wt - | mol wt ~44 kDa | mol wt - |
| solubility 0.1 M phosphate buffer: soluble (pH 6.0), H2O: soluble | solubility 50 mM potassium phosphate buffer, pH 6.0: 5 mg/mL (0.1% BSA) | solubility - | solubility 0.1 M phosphate, pH 6.0-7.0: 5 mg/mL |
| Quality Level 200 | Quality Level 100 | Quality Level 200 | Quality Level 100 |
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signalword
Danger
hcodes
pcodes
Hazard Classifications
Resp. Sens. 1
Storage Class
11 - Combustible Solids
wgk
WGK 1
ppe
Eyeshields, Gloves, type N95 (US)
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