Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704) couples the oxidation of NADPH by cytochrome P450 reductase (CPR) to the reduction of a nearly colorless probe into a brightly colored product with an absorbance peak at OD=460 nm, with the rate of color generation being directly proportional to CPR activity.
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- Stem cell reports 17:2050-20632022P450 oxidoreductase regulates barrier maturation by mediating retinoic acid metabolism in a model of the human BBB.Applications:Unspecified applicationReactive species:Unspecified reactive speciesDor Zlotnik,Tatiana Rabinski,Aviv Halfon,Shira Anzi,Inbar Plaschkes,Hadar Benyamini,Yuval Nevo,Orly Yahalom Gershoni,Benyamin Rosental,Eli Hershkovitz,Ayal Ben-Zvi,Gad D VatinePubMed 35961311
Key facts
- Detection method
Colorimetric
- Sample types
Purified protein, Tissue Lysate, Microsomes, Cell Lysate
- Assay type
Enzyme activity
- Reactive species
Mammals
- Sensitivity
= 0.2 mU/well
Target data
Function
This enzyme is required for electron transfer from NADP to cytochrome P450 in microsomes. It can also provide electron transfer to heme oxygenase and cytochrome B5.
Alternative names
NADPH--cytochrome P450 reductase, CPR, P450R, POR, CYPOR
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Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704) couples the oxidation of NADPH by cytochrome P450 reductase (CPR) to the reduction of a nearly colorless probe into a brightly colored product with an absorbance peak at OD=460 nm, with the rate of color generation being directly proportional to CPR activity.
Alternative names
NADPH--cytochrome P450 reductase, CPR, P450R, POR, CYPOR
Key facts
- Detection method
Colorimetric
- Sample types
Purified protein, Tissue Lysate, Microsomes, Cell Lysate
- Assay type
Enzyme activity
- Reactive species
Mammals
- Assay Platform
Microplate reader
- Sensitivity
= 0.2 mU/well
Storage
- Shipped at conditions
Blue Ice
- Appropriate short-term storage conditions
-20°C
- Appropriate long-term storage conditions
-20°C
- Storage information
-20°C
Notes
Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704) couples the oxidation of NADPH by cytochrome P450 reductase (CPR) to the reduction of a nearly colorless probe into a brightly colored product with an absorbance peak at OD=460 nm, with the rate of color generation being directly proportional to CPR activity. The NADPH utilized by CPR is generated in situ from β-NADP+ via oxidation of glucose-6-phosphate (G6P) to 6-phospho-D-glucono-1,5-lactone by glucose-6-phosphatase dehydrogenase (G6PDH). The kit can be used to determine CPR activity in a variety of samples, with a detection limit of ~0.2 mU of CPR activity per reaction.
For assessment of CPR activity in crude biological samples that may have extraneous reductases capable of reducing the substrate, an inhibitor of NADPH-dependent flavoproteins is included. In this case, the specific CPR activity may be calculated by running parallel reactions in the presence and absence of inhibitor and subtracting any residual activity detected with the inhibitor present. The kit contains sufficient reagents for performing 100 reactions in a 96-well plate format.
This product is manufactured by BioVision, an Abcam company and was previously called K700 Cytochrome P450 Reductase (CPR) Activity Kit (Colorimetric). K700-100 is the same size as the 100 test size of ab204704.
NADPH-cytochrome P450 reductase (CPR, EC 1.6.2.4) is a ~78 kDa membrane-bound flavoenzyme that catalyzes the transfer of electrons from NADPH to members of the cytochrome P450 monooxidase (CYP) enzyme family in the endoplasmic reticulum. CPR contains two tightly bound flavin cofactors, FAD and FMN, which participate in the sequential transfer of electrons from NADPH→FAD→FMN→CYP, oxidizing NADPH to NADP+ and reducing the CYP heme moiety to the substrate- and oxygen-binding ferrous state. As CPR is required for the function of all CYP isozymes, it plays a critical role in the metabolism of drugs, organic pollutants and other xenobiotic compounds, in addition to its role in biosynthesis of certain vitamins and steroid hormones.
Supplementary info
This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
Cytochrome P450 reductase also known as NADPH--cytochrome P450 reductase (CPR) is an enzyme important in electron transfer. It has a mass of approximately 77 kDa and is responsible for transferring electrons from NADPH to cytochrome P450 enzymes supporting their function. The enzyme is predominantly expressed in the endoplasmic reticulum of liver cells and other tissues where detoxification processes occur. Cytochrome reductase as it is sometimes called plays a fundamental role in the metabolism of drugs and the synthesis of cholesterol steroids and other lipids.
- Biological function summary
CPR facilitates electron donation to all microsomal cytochrome P450 enzymes which are involved in a variety of oxidative reactions. This protein is essential for P450 activity assay as it is part of a multienzyme complex that metabolizes exogenous and endogenous compounds efficiently. Without cytochrome P450 reductase the catalytic activity of cytochrome P450 enzymes becomes impaired affecting metabolizing mechanisms for various substances and having systemic biological impacts.
- Pathways
Cytochrome P450 reductase directly participates in drug metabolism and steroid biosynthesis pathways. It together with the cytochrome P450 enzymes metabolizes drugs to make them more water-soluble for excretion. The proteins 3 beta-hydroxysteroid dehydrogenase and CYP3A4 are some of the related components within these pathways working in conjunction to achieve metabolic homeostasis. P450 reductase assays are often utilized to study these pathways further understanding the biological roles and interactions of these proteins.
- Associated diseases and disorders
CPR has a significant connection to conditions like congenital adrenal hyperplasia and prostate cancer. Mutations or dysregulations in cytochrome P450 reductase can lead to accumulated steroid precursors and disrupted steroid hormone synthesis contributing to these disorders. The relationship with adrenal enzyme 21-hydroxylase and the androgen receptor illustrates the pathway complexities and the clinical relevance of cytochrome P450 reductase. Understanding these connections highlights the importance of CPR in maintaining normal physiological functions and how its abnormalities can lead to serious health issues.
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3 product images
Functional Studies - Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704)
Typical G6P Standard curve using Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704). One mole G6P corresponds to one mole of β-NADP+ reduced to NADPH, which subsequently generates one mole of reduced substrate.
Functional Studies - Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704)
Reaction kinetics of recombinant human CPR (positive control) and rat microsomal CPR (with and without inhibitor).
Functional Studies - Cytochrome P450 Reductase Activity Assay Kit (Colorimetric) (ab204704)
Relative CPR activity detected in rat liver microsomes (RLM, 25 μg total protein) and HepG2 cell lysate (40 μg total protein).
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