P6140

Peroxidase aus Meerrettich

Type X, ammonium sulfate suspension

• Synonym(e): Donor:hydrogen-peroxide oxidoreductase, Meerrettichperoxidase
• CAS-Nummer: 9003-99-0
• eCl@ss: 32160410
• UNSPSC Code: 12352204
• NACRES: NA.54
• EC Number: 232-668-6
• MDL number: MFCD00071339
• EG-Nummer: 1.11.1.7 (BRENDA, IUBMB)
• Specific activity: ≥225 units/mg protein (biuret, using pyrogallol)
• Biological source: horseradish
• Concentration: ≥1.0 mg/mL

Eigenschaften

• biological source: horseradish
• Quality Segment: 200
• type: Type X
• form: ammonium sulfate suspension
• specific activity: ≥225 units/mg protein (biuret, using pyrogallol)
• mol wt: ~44 kDa
• concentration: ≥1.0 mg/mL
• absorbance ratio: RZ 2.5-3.5
• storage temp.: 2-8°C
• SMILES string: [O+H2]O[O-]
• InChI: 1S/H2O3/c1-3-2/h1-2H
• InChI key: JSPLKZUTYZBBKA-UHFFFAOYSA-N

Beschreibung

General description

Horseradish peroxidase (HRP) wird aus Meerrettich (Amoracia rusticana) Wurzeln isoliert und ist innerhalb der Peroxidasen der Gruppe der Ferroprotoporpyhrine zuzuordnen. HRP besteht aus einer einzelnen Polypeptidkette und entält vier Disulfidbrücken. Sie ist ein Glykoprotein mit einem Kohlenhydratanteil von 18%. Der Kohlenhydratanteil besteht aus Galactose, Arabinose, Xylose, Fucose, Mannose, Mannosamin und Galactosamin - in Mengenverhältnissen abhängig vom Isoenzym. Ihr Molekulargewicht (~44 kDa) setzt sich aus der Polypeptidkette (33,890 Daltons), dem Hemin plus Ca²⁺ (~700 Daltons) und der Glykosylierung (~9,400 Daltons) zusammen. Es existieren zumindest sieben Isoenzyme von HRP. Der isoelektrische Punkt von HRP ist in einem Bereich von pH 3 3.0 - 9.0.

Application

Horseradish peroxidase (HRP) is isolated from horseradish roots (Amoracia rusticana). 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[1]. Product P6140 has been used to detect low density lipoprotein (LDL)[2].

The enzyme from Sigma has been used while assessing the skin sensitization potential of pro-haptens.[3] It has also been used to show that peroxidase (PO) activity and its heat stability correlate with the availability of free Ca²⁺ ions.[4]

Biochem/physiol Actions

When incubated with a substrate, horseradish peroxidase produces a coloured, fluorimetric, or luminescent derivative of the labeled molecule, allowing quantification[1]. Horseradish peroxidase has been shown to slightly reduce the level of inhibition in a cydAB mutant[5].

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 that include the use of glutaraldehyde, periodate oxidation, 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. 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. Sodium azide, cyanide, L-cystine, dichromate, ethylenethiourea, hydroxylamine, sulfide, vanadate, p-aminobenzoic acid, and Cd²⁺, Co²⁺, Cu²⁺, Fe³⁺, Mn²⁺, Ni²⁺, Pb²⁺ ions are found to inhibit its enzyme activity.

Physical form

Crystalline suspension in 3.2 M (NH₄)₂SO₄ solution containing potassium phosphate buffer, pH 6.0

Preparation Note

Water may be used to dilute suspension if needed.

Analysis Note

Die RZ (Reinheitszahl) ist das Absorptionsratio A₄₀₃/A₂₇₅ bestimmt bei 0.5-1.0 mg/ml in deionisiertem Wasser. Es ist ein Mass für den Hämingehalt, nicht für die Enzymaktivität. Selbst Zubereitungen mit hoher RZ können eine tiefe Enzymaktivität haben.

Preliminary studies indicate the presence of two basic and no acidic isoenzymes

Other Notes

Eine Pyrogalloleinheit bildet in 20 s bei einem pH-Wert von 6,0 und 20 °C 1 mg Purpurogallin aus Pyrogallol.

Weitere Informationen über Peroxidase finden Sie unter www.sigma-aldrich.com/enzymeexplorer.

Sicherheitshinweise

• pictograms: GHS08
• signalword: Danger
• hcodes: H334
• pcodes: P261 - P284 - P501
• Hazard Classifications: Resp. Sens. 1
• Lagerklasse: 11 - Combustible Solids
• wgk: WGK 1
• ppe: Eyeshields, Gloves, type N95 (US)