GO:0044535 very-long-chain fatty acyl-CoA oxidase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0044535 describes the peroxisomal enzyme activity that catalyzes the first step of very-long-chain fatty acid beta-oxidation, converting a very-long-chain 2,3-saturated fatty acyl-CoA to a very-long-chain (2E)-enoyl-CoA with release of H2O2.
• The reaction is carried out by acyl-CoA oxidase enzymes, principally ACOX1 in mammals, which initiate peroxisomal beta-oxidation of substrates that cannot be handled efficiently by mitochondria [2,5].
• Very-long-chain fatty acyl-CoA oxidase activity is central to lipid homeostasis; loss or dysregulation causes accumulation of very-long-chain fatty acids and contributes to steatohepatitis, metabolic disease, and cancer biology [1,3,5].
• ACOX1-dependent very-long-chain fatty acyl-CoA oxidase activity influences circulating lipids and adipose remodeling, linking hepatic peroxisomal oxidation to systemic metabolic health.
• In multiple myeloma, the ABCD1-ACOX1 axis supports tumor growth and can be targeted to suppress disease, showing that this activity has oncogenic roles beyond fatty acid catabolism.
• Studying GO:0044535 requires combining enzyme assays, organelle biology, and CRISPR models because the activity is compartmentalized in peroxisomes and coupled to H2O2 production [2,6].
Description
Very-long-chain fatty acyl-CoA oxidase activity (GO:0044535) is a molecular function that catalyzes the reaction: a very-long-chain 2,3-saturated fatty acyl-CoA + O2 = a very-long-chain (2E)-enoyl-CoA + H2O2. This activity is the rate-limiting entry step of peroxisomal beta-oxidation for fatty acids that are too long to be efficiently oxidized by mitochondria, and it is therefore essential for lipid homeostasis [2,5]. In mammals, the enzyme ACOX1 is the principal catalyst of this activity, and its dysfunction leads to accumulation of very-long-chain fatty acids and downstream pathology [1,5]. For researchers, GO:0044535 matters because it sits at the intersection of peroxisomal biology, oxidative stress, and metabolic disease. The reaction directly generates hydrogen peroxide, so changes in flux through this step alter cellular redox balance and signaling [6,7]. Recent work shows that hepatic ACOX1 activity regulates circulating lipids and adipose remodeling, indicating systemic metabolic consequences of this single enzymatic step. In cancer, ACOX1-dependent very-long-chain fatty acyl-CoA oxidase activity supports tumor growth in multiple myeloma and is linked to metabolic reprogramming in hepatocellular carcinoma [3,4]. This article provides a research-grade overview of GO:0044535, covering its definition, mechanism, key genes, disease relevance, and the experimental models, including CRISPR knockout, point mutation, knock-in, and overexpression, that are used to study it.
very-long-chain fatty acyl-CoA oxidase activity At A Glance
| GO ID | GO:0044535 |
|---|---|
| GO term | very-long-chain fatty acyl-CoA oxidase activity |
| Ontology | molecular_function |
| Synonym | very-long-chain acyl-CoA oxidase activity; very long chain fatty-acyl-CoA oxidase activity; VLC fatty-acyl-CoA oxidase activity |
| Definition | Catalysis of the reaction: a very-long-chain 2,3-saturated fatty acyl-CoA + O2 = a very-long-chain (2E)-enoyl-CoA + H2O2 |
| Major function | Initiates peroxisomal beta-oxidation of very-long-chain fatty acids |
| Representative enzyme | ACOX1 (acyl-CoA oxidase 1) in mammals |
| Subcellular location | Peroxisome |
| Reaction product | Very-long-chain (2E)-enoyl-CoA and hydrogen peroxide |
What Is GO:0044535?
GO:0044535, very-long-chain fatty acyl-CoA oxidase activity, is defined as the catalysis of the reaction in which a very-long-chain 2,3-saturated fatty acyl-CoA is oxidized by molecular oxygen to form a very-long-chain (2E)-enoyl-CoA and hydrogen peroxide. In other words, it is the peroxisomal oxidase step that desaturates very-long-chain fatty acyl-CoA substrates, introducing a trans double bond at the 2,3 position while producing H2O2 [2,6]. This activity is distinguished from mitochondrial acyl-CoA dehydrogenases by its use of O2 as the electron acceptor and by its preference for very-long-chain substrates [2,7].
Why Is very-long-chain fatty acyl-CoA oxidase activity Important in Cell Biology?
Very-long-chain fatty acyl-CoA oxidase activity is important because it controls the first committed step of peroxisomal beta-oxidation for fatty acids that cannot be oxidized in mitochondria, and its dysfunction causes very-long-chain fatty acid accumulation, oxidative stress, and metabolic disease [2,5]. Because the reaction produces H2O2, it also directly contributes to cellular redox signaling and damage [6,7]. In addition, this activity has emerged as a metabolic vulnerability in cancer, where ACOX1-dependent oxidation supports tumor growth and can be targeted therapeutically [3,4].
• Initiates peroxisomal beta-oxidation of very-long-chain fatty acids, a pathway essential for lipid homeostasis.
• Prevents toxic accumulation of very-long-chain fatty acids that occurs when peroxisomal oxidation is impaired.
• Generates hydrogen peroxide, linking fatty acid oxidation to oxidative stress and redox signaling [6,7].
• Regulates circulating lipid levels and adipose tissue remodeling through hepatic ACOX1 activity.
• Supports tumor growth in multiple myeloma via the ABCD1-ACOX1-MET/IGF1R axis.
• Is reprogrammed in hepatocellular carcinoma in association with mitochondrial fission and SIRT1 suppression.
• Provides a mechanistic explanation for steatohepatitis associated with peroxisomal beta-oxidation defects.
• Represents a potential therapeutic target for metabolic disorders and cancers dependent on fatty acid oxidation [1,4].
• Serves as a biomarker of peroxisomal function in inherited metabolic myopathies.
• Enables researchers to dissect peroxisome-specific versus mitochondrial fatty acid oxidation using selective substrates and inhibitors.
What Happens During very-long-chain fatty acyl-CoA oxidase activity?
Substrate recognition and binding
In simple terms: The enzyme first grabs a very-long-chain fatty acyl-CoA molecule and holds it in place.
Very-long-chain fatty acyl-CoA oxidase activity begins with the binding of a very-long-chain 2,3-saturated fatty acyl-CoA substrate to the enzyme active site. The enzyme shows preference for substrates with chain lengths that are poorly handled by mitochondrial beta-oxidation, which is why this activity is compartmentalized in peroxisomes [2,5]. Substrate binding positions the fatty acyl chain for oxidation at the C2-C3 bond.
Oxidative desaturation and H2O2 release
In simple terms: Oxygen is used to remove hydrogen from the fatty acid, creating a double bond and releasing hydrogen peroxide.
The catalytic step transfers electrons from the fatty acyl-CoA substrate to molecular oxygen, forming a very-long-chain (2E)-enoyl-CoA and hydrogen peroxide. This reaction is characteristic of acyl-CoA oxidases, which use O2 rather than the electron-transferring flavoprotein used by mitochondrial acyl-CoA dehydrogenases [2,7]. The H2O2 produced is a reactive oxygen species that can be detoxified by peroxisomal catalase or contribute to redox signaling.
Coupling to downstream beta-oxidation
In simple terms: The product of this step is passed down a chain of reactions that shortens the fatty acid.
The very-long-chain (2E)-enoyl-CoA produced by GO:0044535 is further processed by the remaining enzymes of peroxisomal beta-oxidation, including enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, and 3-ketoacyl-CoA thiolase. Repeated cycles shorten the fatty acid chain, after which the shortened products can be exported to mitochondria for complete oxidation [2,5]. Thus, very-long-chain fatty acyl-CoA oxidase activity is the rate-limiting entry point for peroxisomal fatty acid chain shortening.
Regulation by substrate availability and enzyme abundance
In simple terms: How fast this reaction goes depends on how much substrate is available and how much enzyme is present.
Flux through very-long-chain fatty acyl-CoA oxidase activity is influenced by the availability of very-long-chain fatty acyl-CoA substrates and by the expression level of the oxidase enzyme, principally ACOX1 in mammals [1,2]. Hepatic ACOX1 levels are regulated in response to metabolic state and influence circulating lipids and adipose remodeling. In cancer cells, ACOX1 expression and activity can be reprogrammed to support fatty acid oxidation and tumor growth [3,4].
Key Genes Involved in GO:0044535 very-long-chain fatty acyl-CoA oxidase activity
The following genes and proteins are directly or functionally linked to very-long-chain fatty acyl-CoA oxidase activity (GO:0044535) based on published literature.
| Gene | Major Role | Research Relevance |
|---|---|---|
| ACOX1 | Principal mammalian enzyme catalyzing very-long-chain fatty acyl-CoA oxidase activity | Central to peroxisomal beta-oxidation; regulates circulating lipids and adipose remodeling |
| ABCD1 | Peroxisomal transporter importing very-long-chain fatty acyl-CoA substrates | Supports ACOX1-dependent oxidation; targeting ABCD1-ACOX1 axis suppresses multiple myeloma |
| CAT | Peroxisomal catalase detoxifying H2O2 produced by the oxidase reaction | Modulates oxidative stress downstream of very-long-chain fatty acyl-CoA oxidase activity |
| SIRT1 | Deacetylase linked to mitochondrial dynamics and fatty acid metabolism | Its suppression is associated with reprogrammed fatty acid metabolism in hepatocellular carcinoma |
| MET | Receptor tyrosine kinase downstream of ABCD1-ACOX1 signaling | Part of the ABCD1-ACOX1-MET/IGF1R axis in multiple myeloma |
| IGF1R | Receptor tyrosine kinase cooperating with MET in oncogenic signaling | Component of the ABCD1-ACOX1-MET/IGF1R axis in multiple myeloma |
| ACOX2 | Related acyl-CoA oxidase with distinct substrate specificity | Provides comparative context for peroxisomal oxidase family function |
| ACOX3 | Related acyl-CoA oxidase involved in peroxisomal beta-oxidation | Helps distinguish very-long-chain specific activity from other oxidase activities |
| HADHA | Mitochondrial trifunctional protein subunit for long-chain fatty acid oxidation | Contrasts mitochondrial versus peroxisomal oxidation pathways |
| CPT1A | Mitochondrial outer membrane carnitine palmitoyltransferase | Defines the boundary between mitochondrial and peroxisomal fatty acid oxidation |
| PPARA | Nuclear receptor regulating lipid metabolism genes | Controls expression of fatty acid oxidation genes including ACOX1 |
| NR1H3 | Liver X receptor alpha regulating lipid homeostasis | Linked to hepatic lipid handling and ACOX1-dependent pathways |
| ETFA | Electron transfer flavoprotein subunit for mitochondrial dehydrogenases | Highlights the different electron acceptor used by oxidases versus dehydrogenases |
| SCP2 | Peroxisomal sterol carrier protein involved in lipid transfer | Supports peroxisomal lipid metabolism context |
| PEX5 | Peroxisomal targeting signal receptor | Required for import of ACOX1 into peroxisomes |
| PEX7 | Peroxisomal targeting signal 2 receptor | Contributes to peroxisomal protein import relevant to oxidase function |
| GNPAT | Peroxisomal enzyme in ether lipid synthesis | Illustrates broader peroxisomal metabolic context |
| FAR1 | Fatty acyl-CoA reductase involved in very-long-chain fatty acid metabolism | Relevant to substrate supply for very-long-chain fatty acyl-CoA oxidase activity |
How Is very-long-chain fatty acyl-CoA oxidase activity Regulated?
Very-long-chain fatty acyl-CoA oxidase activity is regulated at multiple levels. Substrate supply depends on peroxisomal import of very-long-chain fatty acyl-CoAs, which involves ABCD1 and other transporters. Enzyme abundance is controlled transcriptionally by lipid-sensing nuclear receptors such as PPARA, which induces fatty acid oxidation genes including ACOX1. In cancer cells, ACOX1 expression and activity can be reprogrammed in response to metabolic stress and signaling changes, as seen in hepatocellular carcinoma where increased mitochondrial fission and SIRT1 suppression accompany altered fatty acid metabolism. The H2O2 produced by the reaction is itself a regulatory signal that can influence redox-sensitive pathways and is balanced by peroxisomal catalase [6,7].
very-long-chain fatty acyl-CoA oxidase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| ACOX1 | Metabolic disease, steatohepatitis, circulating lipid regulation | Liver-specific Acox1 knockout mouse; ACOX1 overexpression hepatocytes [1,5] |
| ABCD1 | Multiple myeloma, peroxisomal fatty acid import | ABCD1 knockout myeloma cell lines; xenograft models |
| SIRT1 | Hepatocellular carcinoma metabolic reprogramming | SIRT1 knockout or overexpression HCC cells |
| CAT | Oxidative stress from H2O2 produced by oxidase activity | Catalase knockout cells; H2O2 imaging |
| ACOX1 | Neonatal metabolic myopathy and peroxisomal beta-oxidation defects | Patient-derived fibroblasts; ACOX1 point-mutation knock-in cells |
Metabolic disease and steatohepatitis
Impaired peroxisomal beta-oxidation, including reduced very-long-chain fatty acyl-CoA oxidase activity, leads to accumulation of very-long-chain fatty acids and is associated with steatohepatitis and metabolic dysfunction. Hepatic ACOX1 activity regulates circulating lipid levels and adipose remodeling, so changes in this activity can influence systemic metabolic health. These findings position GO:0044535 as a potential node for therapeutic intervention in metabolic disease [1,5].
Cancer metabolism and multiple myeloma
The ABCD1-ACOX1-MET/IGF1R axis supports multiple myeloma growth, and targeting this axis suppresses disease, demonstrating that very-long-chain fatty acyl-CoA oxidase activity can be oncogenic. In hepatocellular carcinoma, increased mitochondrial fission drives reprogramming of fatty acid metabolism through suppression of SIRT1, linking this activity to tumor metabolic adaptation. These studies suggest that cancers dependent on peroxisomal fatty acid oxidation may be vulnerable to inhibition of GO:0044535 [3,4].
Oxidative stress and inherited metabolic myopathies
Because very-long-chain fatty acyl-CoA oxidase activity produces H2O2, its dysfunction can contribute to oxidative stress in peroxisomal disorders [6,7]. Neonatal metabolic myopathies include disorders of fatty acid oxidation that present with severe energy failure, and peroxisomal beta-oxidation defects are part of this differential diagnosis. Understanding GO:0044535 helps interpret biochemical and genetic findings in these inherited conditions [2,8].
From very-long-chain fatty acyl-CoA oxidase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of ACOX1 abolish very-long-chain fatty acyl-CoA oxidase activity? | ACOX1 knockout cell line or mouse |
| Does a specific ACOX1 missense variant impair catalysis? | ACOX1 point-mutation knock-in cells |
| Can tagged ACOX1 be used to track peroxisomal localization? | Tagged ACOX1 knock-in cells |
| Does ACOX1 overexpression increase very-long-chain fatty acid oxidation? | ACOX1 overexpression cell line |
| Does ABCD1-ACOX1 axis inhibition suppress myeloma growth? | ABCD1 or ACOX1 knockout myeloma xenografts |
| Does altered ACOX1 activity change H2O2 levels? | ACOX1 knockout or overexpression cells with H2O2 sensors |
How to Study the very-long-chain fatty acyl-CoA oxidase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Acyl-CoA oxidase activity assay | Catalytic conversion of very-long-chain fatty acyl-CoA to enoyl-CoA with H2O2 production | Confirming loss or gain of GO:0044535 activity |
| Lipidomics | Very-long-chain fatty acid and acyl-CoA levels | Detecting substrate accumulation after ACOX1 perturbation [1,5] |
| H2O2 imaging | Cellular hydrogen peroxide levels | Linking oxidase activity to redox signaling |
| RNA sequencing | Transcriptional changes in lipid metabolism genes | Identifying pathways downstream of ACOX1 [1,3] |
| Proteomics | Protein abundance and interactions | Characterizing peroxisomal protein networks |
| Immunofluorescence | Peroxisomal localization of ACOX1 | Validating organelle targeting |
| CRISPR knockout screening | Genes required for very-long-chain fatty acid oxidation | Discovering modifiers of GO:0044535 [3,4] |
| Metabolic flux analysis | Flux through fatty acid oxidation pathways | Quantifying pathway activity in cancer and metabolic models [1,4] |
Enzyme activity assays
Very-long-chain fatty acyl-CoA oxidase activity can be measured using spectrophotometric or fluorometric assays that couple the oxidation of very-long-chain fatty acyl-CoA substrates to H2O2 detection [2,6]. These assays are essential to confirm that a genetic perturbation, such as ACOX1 knockout, actually changes catalytic activity rather than only protein abundance [1,2].
Lipidomics and fatty acid profiling
Because GO:0044535 controls very-long-chain fatty acid catabolism, mass spectrometry-based lipidomics can quantify accumulation of very-long-chain fatty acids in cells or tissues with altered enzyme activity [1,5]. Such profiling links molecular function to metabolic phenotypes and disease states [1,5].
Imaging of peroxisomes and H2O2
Fluorescent reporters for peroxisomes and genetically encoded H2O2 sensors allow researchers to visualize where very-long-chain fatty acyl-CoA oxidase activity occurs and how it affects local redox state [6,7]. These methods complement biochemical assays by providing spatial information [2,6].
Transcriptomics and proteomics
RNA sequencing and proteomics can reveal how perturbation of very-long-chain fatty acyl-CoA oxidase activity reshapes lipid metabolic gene networks, as shown in studies of ACOX1-dependent metabolic remodeling [1,3]. These approaches help identify downstream pathways and potential therapeutic targets [1,3].
How CRISPR Can Be Used to Study GO:0044535 very-long-chain fatty acyl-CoA oxidase activity
Knockout
CRISPR knockout of ACOX1 is used to eliminate very-long-chain fatty acyl-CoA oxidase activity and study consequences for lipid accumulation, redox balance, and disease phenotypes [1,2]. Knockout models help determine whether a phenotype depends specifically on this enzymatic activity.
Point Mutation
Point-mutation knock-in can model disease-associated ACOX1 variants or catalytically dead enzymes, allowing separation of catalytic activity from other protein functions. Such models are valuable for testing whether specific residues are required for very-long-chain fatty acyl-CoA oxidase activity [2,8].
Knock-in
Tagged knock-in of ACOX1 enables tracking of enzyme localization and interactions in peroxisomes without overexpression artifacts. Knock-in of reporter cassettes can also be used to monitor pathway activity in live cells.
Overexpression
Overexpression of ACOX1 increases very-long-chain fatty acyl-CoA oxidase activity and can drive metabolic remodeling, as shown in studies of hepatic ACOX1 and circulating lipids. Overexpression models are useful for testing sufficiency of the activity in disease phenotypes [1,3].
How EDITGENE Supports very-long-chain fatty acyl-CoA oxidase activity Research
Researchers studying very-long-chain fatty acyl-CoA oxidase activity-related genes often need to determine whether a candidate gene is causally involved in peroxisomal fatty acid oxidation, metabolic disease, or cancer. EDITGENE provides CRISPR-based cell models and screening services to test these hypotheses rigorously.
Contact EDITGENE today to design your custom CRISPR model for very-long-chain fatty acyl-CoA oxidase activity research.
Frequently Asked Questions About very-long-chain fatty acyl-CoA oxidase activity
What is very-long-chain fatty acyl-CoA oxidase activity?
It is the enzyme activity defined by GO:0044535 that catalyzes the conversion of a very-long-chain 2,3-saturated fatty acyl-CoA to a very-long-chain (2E)-enoyl-CoA with release of H2O2.
What genes are involved in very-long-chain fatty acyl-CoA oxidase activity?
The principal gene is ACOX1, which encodes the mammalian acyl-CoA oxidase that carries out this activity; ABCD1 supplies substrates, and CAT detoxifies the H2O2 produced [1,2,4].
What is the GO ID for very-long-chain fatty acyl-CoA oxidase activity?
The GO ID is GO:0044535, and the ontology aspect is molecular_function.
Where does very-long-chain fatty acyl-CoA oxidase activity occur in the cell?
It occurs in peroxisomes, where very-long-chain fatty acids undergo beta-oxidation [2,5].
Why is very-long-chain fatty acyl-CoA oxidase activity important in disease?
Its dysfunction causes very-long-chain fatty acid accumulation and is linked to steatohepatitis, metabolic disease, and cancer growth, including multiple myeloma and hepatocellular carcinoma [1,3,4,5].
How is very-long-chain fatty acyl-CoA oxidase activity measured?
It is measured using enzyme assays that detect substrate conversion or H2O2 production, often combined with lipidomics and imaging [2,6].
What happens when ACOX1 is knocked out?
ACOX1 knockout eliminates very-long-chain fatty acyl-CoA oxidase activity, leading to accumulation of very-long-chain fatty acids and altered lipid metabolism [1,2].
Is very-long-chain fatty acyl-CoA oxidase activity involved in cancer?
Yes, ACOX1-dependent activity supports multiple myeloma growth through the ABCD1-ACOX1-MET/IGF1R axis and is reprogrammed in hepatocellular carcinoma [3,4].
What is the difference between very-long-chain fatty acyl-CoA oxidase and mitochondrial acyl-CoA dehydrogenase?
The oxidase uses O2 and produces H2O2 in peroxisomes, whereas mitochondrial dehydrogenases transfer electrons to the electron transfer flavoprotein [2,7].
How can CRISPR help study very-long-chain fatty acyl-CoA oxidase activity?
CRISPR knockout, point mutation, knock-in, and overexpression models allow researchers to test causality and dissect the role of GO:0044535 in disease [1,4,8].
Conclusion
Very-long-chain fatty acyl-CoA oxidase activity (GO:0044535) is a peroxisomal molecular function that initiates beta-oxidation of very-long-chain fatty acids and produces H2O2, making it central to lipid homeostasis and redox biology [2,6]. Its dysregulation is linked to metabolic disease, steatohepatitis, and cancer, and ACOX1 is the principal enzyme responsible in mammals [1,3,4,5]. Researchers can now use CRISPR knockout, point mutation, knock-in, overexpression, and library screening to dissect the causal roles of ACOX1 and related genes in health and disease, with EDITGENE providing end-to-end model generation and bioinformatics support [1,4,8].
References
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- 3. Wu D et al.. 2022. Increased mitochondrial fission drives the reprogramming of fatty acid metabolism in hepatocellular carcinoma cells through suppression of Sirtuin 1.. Cancer Commun (Lond) 42(1):37-55 PMID: 34981667
- 4. Han Z et al.. 2025. Targeting ABCD1-ACOX1-MET/IGF1R axis suppresses multiple myeloma.. Leukemia 39(3):720-733 PMID: 39885295
- 5. Rao MS et al.. 2001. Peroxisomal beta-oxidation and steatohepatitis.. Semin Liver Dis 21(1):43-55 PMID: 11296696
- 6. Kakimoto PA et al.. 2015. H2O2 release from the very long chain acyl-CoA dehydrogenase.. Redox Biol 4:375-80 PMID: 25728796
- 7. Zhang Y et al.. 2019. The fatty acid oxidation enzyme long-chain acyl-CoA dehydrogenase can be a source of mitochondrial hydrogen peroxide.. Redox Biol 26:101253 PMID: 31234015
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