ペルオキシダーゼ from horseradish

西洋ワサビペルオキシダ-ゼは西洋ワサビ(Amoracia rusticana)の根から単離された酵素で、ペルオキシダ-ゼのフェロプロトポルフィリン類に属します。HRPは4個のジスルフィド架橋結合を有する1本のポリペプチドです。18%の糖を含む糖タンパク質です。糖の組成はガラクト-ス、アラビノ-ス、キシロ-ス、フコ-ス、マンノ-ス、マンノサミン、ガラクトサミンからなり、個々のアイソザイムによって異なります。 この酵素の分子量(約44 kDa)には、ポリペプチド鎖(33,890 Da)、ヘミン+Ca²⁺(約700 Da)、糖(約9,400 Da)が含まれます。HRPには少なくとも7種類のアイソザイムが存在します。 西洋ワサビペルオキシダ-ゼアイソザイムの等電点は3.0~9.0です。

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.

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.

The enzyme has been used to determine H₂O₂ production in tobacco BY-2 cells using a spectrofluorimetric method.

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.

クロマトグラフィ-精製済み

RZ(Reinheitszahl)値は、0.5~1.0 mg/mL脱イオン水溶液で測定したA₄₀₃/A₂₇₅の吸光度比です。これはヘミン含量を測定するもので、酵素活性を測定するものではありません。RZ値の高い標品でも酵素活性が低いことがあります。

予備的な等電点電気泳動のデ-タでは、主にアイソザイムCからなることが示されています。

1ピロガロールユニットは、pH 6.0、20°Cで 20秒あたり1.0 mgのプルプロガリンをピロガロールから生成する酵素量です。

ペルオキシダ-ゼ Type VI(製品番号 P8375)をさらに精製した製品です。

ペルオキシダーゼに関する詳細情は、www.sigma-aldrich.com/enzymeexplorerをご覧ください。