GO:0050664 oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0050664 describes a molecular function in which NADH or NADPH donates electrons to molecular oxygen, reducing it to water or reactive oxygen species.
• This activity is carried out by flavoenzymes and other redox proteins that contain FMN, FAD, or heme cofactors.
• In plants, NAD(P)H-dependent oxygen reduction is linked to carotenoid desaturation in chromoplast membranes.
• The term is distinct from NADH oxidase (GO:0003954) and NADPH oxidase (GO:0016175) because it covers both NADH and NADPH as electron donors.
• Dysregulation of this activity can alter cellular redox balance and contribute to oxidative stress-related diseases.
• CRISPR knockout, point-mutation, and overexpression models are essential to dissect the physiological roles of these enzymes.
Description
GO:0050664, oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor, is a molecular function defined by the catalysis of a redox reaction in which NADH or NADPH serves as the electron donor and molecular oxygen is the electron acceptor. This activity is fundamental to cellular respiration, oxidative stress responses, and various biosynthetic pathways. Enzymes with this activity are found across all kingdoms of life and often utilize flavin or heme cofactors to transfer electrons from NAD(P)H to oxygen. In plants, a 23-kDa oxygen-evolving-complex-like protein is involved in a respiratory redox pathway that links NAD(P)H oxidation to carotene desaturation in chromoplast membranes. Understanding this activity is crucial for researchers studying energy metabolism, antioxidant defense, and the mechanisms of diseases linked to redox imbalance.
oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor At A Glance
| GO ID | GO:0050664 |
|---|---|
| GO term | oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor |
| Ontology | molecular_function |
| Synonym | oxidoreductase activity, acting on NADH or NADPH, oxygen as acceptor |
| Definition | Catalysis of an oxidation-reduction (redox) reaction in which NADH or NADPH acts as a hydrogen or electron donor and reduces an oxygen molecule. |
| Major function | Electron transfer from NAD(P)H to O2, often producing water or reactive oxygen species. |
| Cofactors | FMN, FAD, heme, or other redox-active prosthetic groups. |
| Subcellular location | Cytosol, mitochondria, plastids, and membranes. |
| Representative enzymes | NADH(NADPH)-cytochrome c reductase (FMN-containing), oxygen-evolving-complex-like proteins. |
What Is GO:0050664?
This GO term describes the catalytic activity of an enzyme that transfers electrons from either NADH or NADPH to molecular oxygen (O2), reducing it. The reaction typically produces water (H2O) or, in some cases, superoxide or hydrogen peroxide, depending on the enzyme. The definition explicitly states that NADH or NADPH acts as the hydrogen or electron donor, and oxygen is the acceptor. This activity is classified under oxidoreductases and is distinguished from similar terms by its broad specificity for both NADH and NADPH.
Why Is oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor Important in Cell Biology?
This activity is central to cellular redox homeostasis, energy production, and the response to oxidative stress. By transferring electrons from NAD(P)H to oxygen, these enzymes help maintain the balance of reducing equivalents and can generate signaling molecules such as reactive oxygen species (ROS). In plants, the activity is linked to carotenoid biosynthesis, which is vital for photosynthesis and photoprotection. In humans, related enzymes are implicated in cancer, neurodegeneration, and metabolic disorders. Thus, studying GO:0050664 provides insights into fundamental biology and potential therapeutic targets.
• Maintains cellular redox balance by oxidizing NAD(P)H and reducing oxygen.
• Contributes to the generation of reactive oxygen species (ROS) for signaling and defense.
• Plays a role in plant carotenoid desaturation and chromoplast development.
• Involved in mitochondrial respiration and energy metabolism.
• Dysregulation can lead to oxidative stress, linked to cancer and neurodegeneration.
• Provides targets for antioxidant therapies and metabolic engineering.
• Enzymes with this activity are used in biotechnological applications, such as biosensors.
• Essential for understanding host-pathogen interactions where ROS is used as a defense mechanism.
What Happens During oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor?
Electron Transfer from NAD(P)H to the Enzyme
In simple terms: The enzyme grabs electrons from NADH or NADPH.
The catalytic cycle begins when NADH or NADPH binds to the enzyme's active site. The reduced nicotinamide ring donates a hydride ion (two electrons and a proton) to the enzyme's prosthetic group, such as FMN or FAD. This step is often rate-limiting and can be regulated by substrate availability.
Reduction of Molecular Oxygen
In simple terms: The enzyme passes the electrons to oxygen, turning it into water or other molecules.
The reduced cofactor then transfers electrons to molecular oxygen. Depending on the enzyme, oxygen can be fully reduced to water (four-electron reduction) or partially reduced to superoxide or hydrogen peroxide (one- or two-electron reduction). This step is critical for determining the product and the potential for ROS generation.
Cofactor Regeneration and Product Release
In simple terms: The enzyme resets itself to start the cycle again.
After oxygen reduction, the enzyme's cofactor returns to its oxidized state, and the products (water or ROS) are released. The enzyme is then ready for another round of catalysis. The overall reaction is: NAD(P)H + H+ + O2 -> NAD(P)+ + H2O (or ROS). The stoichiometry and kinetics vary among enzymes.
Coupling to Cellular Processes
In simple terms: This activity is connected to other cellular jobs, like making pigments.
In plants, the redox activity is coupled to carotene desaturation in chromoplast membranes, where a 23-kDa oxygen-evolving-complex-like protein participates in a respiratory redox pathway. This illustrates how the activity can be integrated into biosynthetic pathways beyond simple detoxification.
Key Genes Involved in GO:0050664 oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor
The following genes and proteins are representative of those that exhibit or are associated with oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor.
| Gene | Major Role | Research Relevance |
|---|---|---|
| CYB5R3 | NADH-cytochrome b5 reductase | Involved in fatty acid desaturation and drug metabolism |
| NDI1 | Internal NADH dehydrogenase | Mitochondrial respiration and ROS production |
| AOX1 | Alternative oxidase | Plant and fungal respiration, bypasses cytochrome pathway |
| NOX1 | NADPH oxidase 1 | ROS generation in signaling and host defense |
| NOX2 | NADPH oxidase 2 | Phagocyte oxidative burst |
| NOX4 | NADPH oxidase 4 | Constitutive ROS production, fibrosis |
| NOX5 | NADPH oxidase 5 | Calcium-dependent ROS generation |
| DUOX1 | Dual oxidase 1 | Thyroid hormone synthesis and mucosal defense |
| DUOX2 | Dual oxidase 2 | Thyroid hormone synthesis |
| NQO1 | NAD(P)H quinone dehydrogenase 1 | Detoxification and antioxidant defense |
| POR | Cytochrome P450 oxidoreductase | Electron transfer to P450 enzymes |
| FDX1 | Ferredoxin 1 | Electron transfer in mitochondria |
| CYPOR | Cytochrome P450 oxidoreductase | Drug metabolism and steroidogenesis |
| MICAL1 | Microtubule associated monooxygenase | Actin dynamics and redox signaling |
| MICAL2 | Microtubule associated monooxygenase | Actin dynamics and redox signaling |
| MSRA | Methionine sulfoxide reductase A | Repair of oxidized methionine |
| MSRB1 | Methionine sulfoxide reductase B1 | Repair of oxidized methionine |
How Is oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor Regulated?
The activity of enzymes with GO:0050664 is regulated at multiple levels. Transcriptional regulation controls enzyme abundance in response to oxidative stress, hypoxia, and metabolic demands. Post-translational modifications, such as phosphorylation, can modulate catalytic efficiency. Allosteric regulation by NAD(P)H/NAD(P)+ ratios and feedback inhibition by products (e.g., ROS) fine-tunes activity. In plants, the 23-kDa oxygen-evolving-complex-like protein is regulated by developmental and environmental cues. Additionally, cofactor availability (FMN, FAD, heme) and subcellular localization influence overall activity.
oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| NOX1 | Colorectal cancer | Knockout in HCT116 cells |
| NOX2 | Chronic granulomatous disease | Point mutation in PLB-985 cells |
| NOX4 | Pulmonary fibrosis | Overexpression in A549 cells |
| NQO1 | Cancer chemoresistance | Knockout in MCF-7 cells |
| MSRA | Neurodegeneration | Knock-in in SH-SY5Y cells |
Cancer
Altered expression of NAD(P)H oxidases and related enzymes can lead to increased ROS production, promoting genomic instability, proliferation, and survival. For example, NOX1 and NOX4 are overexpressed in various cancers and contribute to tumorigenesis. Targeting these enzymes with inhibitors or CRISPR knockout is a promising therapeutic strategy.
Neurodegeneration
Oxidative stress is a hallmark of neurodegenerative diseases such as Alzheimer's and Parkinson's. Dysregulated NAD(P)H-dependent oxygen reduction can exacerbate ROS accumulation, leading to neuronal damage. Enzymes like NQO1 and MSRA play protective roles, and their dysfunction is linked to disease progression.
Metabolic Disorders
Redox imbalance contributes to insulin resistance, obesity, and cardiovascular diseases. NADPH oxidases are implicated in endothelial dysfunction and inflammation. Modulating their activity through genetic models can elucidate their roles in metabolic syndrome.
Infectious Diseases
Phagocytic NOX2 generates ROS to kill pathogens. Defects in NOX2 cause chronic granulomatous disease, characterized by recurrent infections. Studying this activity helps understand host-pathogen interactions and develop immunotherapies.
From oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of NOX1 reduce ROS and tumor growth? | NOX1 knockout in cancer cell lines |
| Does a specific NOX2 mutation impair phagocyte function? | Point mutation knock-in in iPSCs |
| Can overexpression of NQO1 protect against oxidative stress? | NQO1 overexpression in neuronal cells |
| What is the subcellular localization of AOX1? | Tagged knock-in with GFP in plant cells |
| Is the 23-kDa protein essential for carotene desaturation? | Knockout in Narcissus pseudonarcissus chromoplasts |
| Does NOX4 inhibition reverse fibrosis? | Inducible knockout in mouse models |
How to Study the oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor Process
| Method | What It Measures | Typical Application |
|---|---|---|
| NAD(P)H oxidation assay | Enzyme kinetics | Purified enzyme characterization |
| Oxygen consumption assay | O2 consumption rate | Mitochondrial respiration |
| Amplex Red | H2O2 production | Cellular ROS detection |
| DCFDA | Total ROS | High-throughput screening |
| CRISPR knockout | Gene function | Loss-of-function studies |
| RNA-seq | Transcriptional changes | Pathway analysis |
| Redox proteomics | Oxidized proteins | Biomarker discovery |
Enzymatic Activity Assays
Spectrophotometric assays monitor the oxidation of NAD(P)H at 340 nm or the reduction of oxygen using oxygen electrodes. These methods provide kinetic parameters (Km, Vmax) and are essential for characterizing enzymes with GO:0050664.
ROS Detection
Fluorescent probes such as DCFDA, Amplex Red, and MitoSOX measure ROS production in live cells. These assays link enzyme activity to cellular oxidative stress and signaling.
Genetic Knockout and Knockdown
CRISPR/Cas9 knockout or siRNA knockdown of candidate genes followed by phenotypic analysis (e.g., proliferation, ROS levels) helps establish causality. This is particularly useful for distinguishing among NOX family members.
Proteomics and Redox Proteomics
Mass spectrometry-based approaches identify oxidized proteins and post-translational modifications, revealing downstream effects of NAD(P)H-dependent oxygen reduction.
How CRISPR Can Be Used to Study GO:0050664 oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor
Knockout
CRISPR knockout of genes encoding GO:0050664 enzymes (e.g., NOX1, NOX4) abolishes their activity, allowing researchers to study loss-of-function phenotypes such as reduced ROS production, altered cell proliferation, or metabolic reprogramming. This approach is ideal for validating drug targets.
Point Mutation
Introducing specific point mutations (e.g., in the NADPH binding site of NOX2) via CRISPR base editing or HDR can mimic disease-associated variants or dissect catalytic residues. These models help understand structure-function relationships and drug resistance.
Knock-in
Knock-in of tagged versions (e.g., GFP, HA) of these enzymes enables real-time imaging of localization and interaction partners. Knock-in of disease mutations (e.g., in NQO1) creates isogenic models for drug testing.
Overexpression
CRISPR activation (CRISPRa) or lentiviral overexpression of these enzymes increases their activity, useful for studying gain-of-function effects, ROS-mediated signaling, and protection against oxidative stress.
How EDITGENE Supports oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor Research
Researchers studying oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor-related genes often need to determine whether a candidate gene is causally involved in a specific phenotype, such as ROS production or disease progression. EDITGENE provides a comprehensive suite of CRISPR services to create precisely engineered cell models, enabling rigorous functional studies.
Contact EDITGENE today to design your custom CRISPR model for oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor research.
Frequently Asked Questions About oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor
What is GO:0050664?
GO:0050664 is a Gene Ontology molecular function term for oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor. It describes enzymes that transfer electrons from NADH or NADPH to molecular oxygen.
What genes are involved in oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor?
Genes include NOX1, NOX2, NOX4, NQO1, CYB5R3, AOX1, and others that encode enzymes with this activity.
What is the difference between GO:0050664 and NADH oxidase?
GO:0050664 encompasses both NADH and NADPH as electron donors, whereas NADH oxidase (GO:0003954) is specific to NADH.
How is oxidoreductase activity, acting on NAD(P)H, oxygen as acceptor measured?
It is measured using spectrophotometric NAD(P)H oxidation assays, oxygen consumption assays, or ROS detection probes.
What diseases are associated with this activity?
Diseases include cancer, neurodegeneration, chronic granulomatous disease, and metabolic disorders, often due to ROS imbalance.
Can CRISPR be used to study this activity?
Yes, CRISPR knockout, point mutation, knock-in, and overexpression models are powerful tools to dissect gene function in this context.
What cofactors are required for this activity?
Common cofactors include FMN, FAD, and heme, which facilitate electron transfer from NAD(P)H to oxygen.
Is this activity found in plants?
Yes, a 23-kDa oxygen-evolving-complex-like protein in Narcissus pseudonarcissus chromoplasts is involved in a respiratory redox pathway linked to carotene desaturation.
What is the role of this activity in cancer?
It can promote ROS generation, leading to genomic instability and tumor progression; NOX1 and NOX4 are often overexpressed in cancers.
How can EDITGENE help my research?
EDITGENE provides custom CRISPR cell models, library screening, and bioinformatics to study genes with this activity.
Conclusion
GO:0050664 represents a fundamental molecular function that bridges cellular metabolism, redox signaling, and disease. By understanding the enzymes and pathways involved, researchers can develop targeted therapies for oxidative stress-related conditions. CRISPR-based models are indispensable for dissecting these mechanisms, and EDITGENE offers the tools to accelerate discovery.
References
- 1. Johnson MS et al.. 1986. Studies on NADH(NADPH)-cytochrome c reductase (FMN-containing) from yeast: steady-state kinetic properties of the flavoenzyme from top-fermenting ale yeast.. Arch Biochem Biophys 245(1):271-81 PMID: 3080958
- 2. Nievelstein V et al.. 1995. Carotene desaturation is linked to a respiratory redox pathway in Narcissus pseudonarcissus chromoplast membranes. Involvement of a 23-kDa oxygen-evolving-complex-like protein.. Eur J Biochem 233(3):864-72 PMID: 8521852
