GO:0141126 short-chain fatty acyl-CoA hydrolase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0141126 defines the enzymatic activity that hydrolyzes short-chain fatty acyl-CoA thioesters (acyl chains <6 carbons) into free short-chain fatty acids, CoA, and H+.
• This activity is critical for mitochondrial and peroxisomal fatty acid metabolism, controlling the pool of short-chain acyl-CoAs that feed into oxidation, lipogenesis, and signaling.
• Key enzymes include ACOT9, ACOT1/2, and PTE1p, which regulate short-chain fatty acid trafficking and energy homeostasis.
• Dysregulation of short-chain acyl-CoA hydrolases is linked to nonalcoholic fatty liver disease, insulin resistance, and altered lipid metabolism.
• CRISPR knockout, point mutation, and knock-in models are essential to dissect the physiological roles of these hydrolases in vivo.
• Targeting short-chain fatty acyl-CoA hydrolase activity offers therapeutic potential for metabolic disorders and microbial production of short-chain alkanes.
Description
Short-chain fatty acyl-CoA hydrolase activity (GO:0141126) is a molecular function that catalyzes the hydrolysis of short-chain fatty acyl-CoA thioesters, releasing free short-chain fatty acids (SCFAs), coenzyme A (CoA), and a proton. This activity is fundamental to cellular lipid metabolism because it controls the availability of short-chain acyl-CoAs, which are intermediates in fatty acid oxidation, de novo lipogenesis, and protein acylation. Researchers study this term to understand how cells manage energy flux and how disruptions contribute to metabolic diseases such as nonalcoholic fatty liver disease (NAFLD) and insulin resistance. The reaction is conserved from yeast to mammals, with distinct enzymes localized to mitochondria, peroxisomes, and brown adipose tissue. In this article, we integrate authoritative QuickGO annotation with real PubMed literature to provide a comprehensive overview of the mechanism, key genes, disease relevance, and CRISPR-based research methods for GO:0141126.
short-chain fatty acyl-CoA hydrolase activity At A Glance
| GO ID | GO:0141126 |
|---|---|
| GO term | short-chain fatty acyl-CoA hydrolase activity |
| Ontology | molecular_function |
| Synonym | none |
| Definition | Catalysis of the reaction: a short-chain fatty acyl-CoA + H2O = a short fatty acid + CoA + H+. A short-chain fatty acid has an aliphatic tail of less than 6 carbons. |
| Major function | Hydrolysis of short-chain fatty acyl-CoA thioesters to free fatty acids and CoA |
| Substrate specificity | Short-chain fatty acyl-CoAs with acyl chains <6 carbons |
| Localization | Mitochondria, peroxisomes, brown adipose tissue |
| Representative enzymes | ACOT9, ACOT1, ACOT2, PTE1p |
What Is GO:0141126?
GO:0141126 describes the catalytic activity of enzymes that cleave short-chain fatty acyl-CoA molecules in the presence of water, yielding a short-chain fatty acid (aliphatic tail of fewer than 6 carbons), free CoA, and a hydrogen ion. This activity is distinct from medium- and long-chain acyl-CoA hydrolases due to its substrate specificity for short acyl chains. The reaction is a thioester hydrolysis that regulates the cellular levels of short-chain acyl-CoAs, which are key metabolites in mitochondrial and peroxisomal fatty acid degradation and in biosynthetic pathways.
Why Is short-chain fatty acyl-CoA hydrolase activity Important in Cell Biology?
Short-chain fatty acyl-CoA hydrolase activity is essential for maintaining metabolic homeostasis because it controls the intracellular concentrations of short-chain acyl-CoAs, which are pivotal intermediates in fatty acid oxidation, lipogenesis, and signaling. Dysregulation of this activity has been implicated in metabolic disorders such as nonalcoholic fatty liver disease (NAFLD) and insulin resistance, where altered short-chain fatty acid trafficking contributes to hepatic steatosis and glucose dysregulation. Moreover, these enzymes influence the production of short-chain alkanes in microbial systems, highlighting their biotechnological relevance. Understanding GO:0141126 therefore provides insights into both fundamental biochemistry and translational opportunities for metabolic diseases.
• Regulates mitochondrial and peroxisomal fatty acid oxidation by controlling short-chain acyl-CoA levels.
• Modulates de novo lipogenesis and glucose production in the liver.
• Influences energy expenditure in brown adipose tissue through short-chain acyl-CoA hydrolysis.
• Linked to the pathogenesis of nonalcoholic fatty liver disease (NAFLD) and insulin resistance.
• Plays a role in the microbial production of short-chain alkanes, a biofuel precursor.
• Affects protein acetylation and acylation by regulating acetyl-CoA and short-chain acyl-CoA pools.
• Contributes to ketogenic diet responses via mitochondrial acetylation of ACSS1.
• Provides a target for therapeutic intervention in metabolic disorders.
• Essential for efficient degradation of short straight and branched chain fatty acids in yeast.
• Serves as a model for studying thioesterase structure-function relationships.
What Happens During short-chain fatty acyl-CoA hydrolase activity?
Substrate Recognition and Binding
In simple terms: The enzyme grabs a short-chain fatty acyl-CoA molecule.
The first step involves the specific binding of a short-chain fatty acyl-CoA substrate to the active site of the hydrolase. Enzymes such as ACOT9 and PTE1p exhibit preference for acyl chains shorter than six carbons, discriminating against longer chains through steric and hydrophobic interactions. This substrate specificity ensures that only short-chain acyl-CoAs are hydrolyzed, preventing interference with long-chain fatty acid metabolism.
Catalytic Hydrolysis
In simple terms: Water breaks the bond between the fatty acid and CoA.
The catalytic mechanism involves nucleophilic attack by a water molecule on the thioester carbonyl carbon, leading to the cleavage of the thioester bond. This reaction releases a free short-chain fatty acid, CoA, and a proton. The hydrolysis is energetically favorable and does not require ATP, distinguishing it from ligase-mediated activation. The active site typically contains a catalytic triad (serine, histidine, aspartate) that facilitates the hydrolysis.
Product Release and Metabolic Fate
In simple terms: The products are released and used in other pathways.
Following hydrolysis, the free short-chain fatty acid and CoA are released from the enzyme. The fatty acid can then participate in various metabolic pathways, including mitochondrial oxidation, peroxisomal degradation, or export to the cytosol for lipogenesis. CoA is recycled for further acyl-CoA synthesis. In brown adipose tissue, the released fatty acids can be used for thermogenesis.
Integration with Cellular Metabolism
In simple terms: The reaction connects to energy production and fat synthesis.
Short-chain fatty acyl-CoA hydrolase activity is integrated with broader metabolic networks. In the liver, ACOT9 traffics short-chain fatty acids toward de novo lipogenesis and glucose production, linking hydrolysis to hepatic energy homeostasis. In peroxisomes, PTE1p is required for efficient degradation of short straight and branched chain fatty acids, highlighting its role in fatty acid catabolism. This integration ensures that short-chain acyl-CoAs are partitioned appropriately between oxidation and biosynthesis.
Key Genes Involved in GO:0141126 short-chain fatty acyl-CoA hydrolase activity
The following genes encode enzymes with short-chain fatty acyl-CoA hydrolase activity or are directly involved in its regulation, as supported by published literature.
| Gene | Major Role | Research Relevance |
|---|---|---|
| ACOT9 | Mitochondrial acyl-CoA thioesterase that hydrolyzes short-chain acyl-CoAs | Regulates hepatic lipogenesis and glucose production; linked to NAFLD |
| ACOT1 | Cytosolic and peroxisomal acyl-CoA thioesterase with broad specificity | Involved in lipid metabolism and energy homeostasis |
| ACOT2 | Mitochondrial acyl-CoA thioesterase, primarily long-chain but also short-chain | Role in fatty acid oxidation and thermogenesis |
| PTE1p | Peroxisomal acyl-CoA thioesterase in Saccharomyces cerevisiae | Required for degradation of short straight and branched chain fatty acids |
| ACSS1 | Mitochondrial acetyl-CoA synthetase, generates acetyl-CoA for oxidation | Acetylation at K635 alters liver lipid metabolism on ketogenic diet |
| RPN11 | Deubiquitylating enzyme, regulates protein stability | Inhibition ameliorates NAFLD, may affect acyl-CoA hydrolase stability |
| ACOT7 | Brain-specific acyl-CoA thioesterase | May regulate short-chain acyl-CoA in neurons |
| ACOT8 | Peroxisomal acyl-CoA thioesterase | Involved in peroxisomal fatty acid metabolism |
| ACOT11 | Brown adipose tissue acyl-CoA thioesterase | Regulates thermogenesis and energy expenditure |
| ACOT12 | Cytosolic acyl-CoA thioesterase | Modulates acetyl-CoA levels for lipogenesis |
| ACOT13 | Mitochondrial acyl-CoA thioesterase | Role in mitochondrial fatty acid oxidation |
| THEM4 | Acyl-CoA thioesterase with broad specificity | Potential role in lipid signaling |
| THEM5 | Acyl-CoA thioesterase | May hydrolyze short-chain acyl-CoAs |
| BACH1 | Transcription factor regulating oxidative stress | May influence acyl-CoA hydrolase expression |
| SIRT1 | NAD+-dependent deacetylase | Regulates acetylation of metabolic enzymes including ACSS1 |
| PPARA | Nuclear receptor activated by fatty acids | Controls expression of acyl-CoA hydrolases |
| PGC1A | Transcriptional coactivator | Regulates mitochondrial biogenesis and acyl-CoA metabolism |
| UCP1 | Uncoupling protein in brown adipose tissue | Linked to thermogenesis and short-chain acyl-CoA hydrolysis |
How Is short-chain fatty acyl-CoA hydrolase activity Regulated?
Short-chain fatty acyl-CoA hydrolase activity is regulated at multiple levels. Transcriptionally, peroxisome proliferator-activated receptor alpha (PPARA) controls the expression of several acyl-CoA thioesterases in response to fatty acid availability. Post-translationally, acetylation of mitochondrial proteins such as ACSS1 by SIRT1 modulates their activity and affects short-chain acyl-CoA pools. Additionally, the deubiquitylating enzyme RPN11 regulates the stability of metabolic enzymes, and its inhibition ameliorates NAFLD, suggesting a role in the turnover of acyl-CoA hydrolases. Hormonal signals, such as those during ketogenic diet, also influence the acetylation status and activity of these enzymes.
short-chain fatty acyl-CoA hydrolase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| ACOT9 | NAFLD, insulin resistance | Liver-specific knockout mouse, overexpression in hepatocytes |
| RPN11 | NAFLD | RPN11 inhibitor treatment in diet-induced obese mice |
| ACSS1 | Ketogenic diet response, lipid metabolism | ACSS1-K635 acetylation knock-in mouse |
| PTE1p | Fatty acid degradation defects | Yeast pte1 deletion mutant |
| ACOT11 | Obesity, thermogenesis | Brown adipose tissue-specific knockout mouse |
Nonalcoholic Fatty Liver Disease (NAFLD)
Dysregulation of short-chain fatty acyl-CoA hydrolase activity contributes to the pathogenesis of NAFLD. Inhibition of the deubiquitylating enzyme RPN11 ameliorates NAFLD in mice, partly by altering the stability of metabolic enzymes involved in lipid handling. ACOT9, a mitochondrial short-chain acyl-CoA thioesterase, traffics short-chain fatty acids toward de novo lipogenesis and glucose production in the liver, and its dysregulation is associated with hepatic steatosis and insulin resistance. These findings highlight the importance of short-chain acyl-CoA hydrolysis in liver metabolic health.
Insulin Resistance and Type 2 Diabetes
Altered short-chain fatty acid metabolism is linked to insulin resistance. ACOT9-mediated hydrolysis of short-chain acyl-CoAs influences glucose production in the liver, and its overexpression or knockdown affects systemic glucose homeostasis. Additionally, mitochondrial acetylation of ACSS1 on a ketogenic diet alters liver lipid metabolism, which can impact insulin sensitivity. Thus, enzymes with short-chain fatty acyl-CoA hydrolase activity are potential targets for improving insulin sensitivity.
Metabolic Disorders and Thermogenesis
In brown adipose tissue, short-chain acyl-CoA hydrolases regulate thermogenesis by controlling the availability of fatty acids for oxidation. ACOT11 (also known as THEM1) is highly expressed in brown adipose tissue and influences energy expenditure. Dysregulation of these enzymes may contribute to obesity and related metabolic disorders. Furthermore, microbial production of short-chain alkanes, which involves acyl-CoA hydrolysis, has biotechnological implications for biofuel production.
From short-chain fatty acyl-CoA hydrolase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does ACOT9 loss alter hepatic glucose production? | Liver-specific ACOT9 knockout mouse |
| How does ACSS1 acetylation affect ketogenic diet response? | ACSS1-K635Q acetylation mimic knock-in mouse |
| Is PTE1p required for short-chain fatty acid degradation? | Saccharomyces cerevisiae pte1 deletion strain |
| Can RPN11 inhibition treat NAFLD? | RPN11 inhibitor in high-fat diet mouse model |
| What is the role of ACOT11 in thermogenesis? | ACOT11 knockout mouse, cold exposure |
| Can short-chain acyl-CoA hydrolases be targeted for biofuel production? | Engineered microbial strains with acyl-CoA hydrolase overexpression |
How to Study the short-chain fatty acyl-CoA hydrolase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| DTNB assay | CoA release from acyl-CoA | Enzyme kinetics and substrate specificity |
| CRISPR knockout screen | Gene essentiality and modifiers | Identify regulators of ACOT9 expression |
| LC-MS metabolomics | Short-chain acyl-CoA and fatty acid levels | Assess metabolic impact of hydrolase knockout |
| Acetylation proteomics | Post-translational modifications | Study ACSS1 regulation on ketogenic diet |
| RNA-seq | Transcriptional changes | Evaluate PPARA target genes upon hydrolase perturbation |
| Western blot | Protein expression and stability | Assess RPN11 inhibition on acyl-CoA hydrolase levels |
| Yeast growth assay | Fatty acid degradation capacity | Test PTE1p function in Saccharomyces cerevisiae |
| Thermogenesis assay | Oxygen consumption in brown adipocytes | Evaluate ACOT11 role in energy expenditure |
Enzymatic Activity Assays
Direct measurement of short-chain fatty acyl-CoA hydrolase activity is typically performed using spectrophotometric or fluorometric assays that monitor the release of CoA from acyl-CoA substrates. These assays use substrates such as butyryl-CoA or propionyl-CoA and detect the free thiol group of CoA with reagents like DTNB. Such methods are essential for characterizing enzyme kinetics and substrate specificity.
CRISPR-Based Genetic Screens
CRISPR knockout and activation screens can identify genes that regulate short-chain acyl-CoA hydrolase activity. For example, a genome-wide knockout screen in hepatocytes could reveal modifiers of ACOT9 expression or activity. These screens are powerful for discovering novel regulators and potential therapeutic targets.
Metabolomics and Lipidomics
Mass spectrometry-based metabolomics and lipidomics quantify short-chain acyl-CoA and free fatty acid levels in cells and tissues. These methods are used to assess the impact of genetic perturbations (e.g., ACOT9 knockout) on metabolic flux. They provide a systems-level view of how hydrolase activity affects cellular metabolism.
Proteomics and Acetylation Analysis
Proteomic approaches, including immunoprecipitation followed by mass spectrometry, can identify acetylation sites on enzymes like ACSS1 and determine how post-translational modifications regulate their activity. These methods are crucial for understanding the regulatory mechanisms of short-chain acyl-CoA hydrolases.
How CRISPR Can Be Used to Study GO:0141126 short-chain fatty acyl-CoA hydrolase activity
Knockout
CRISPR knockout of genes encoding short-chain fatty acyl-CoA hydrolases (e.g., ACOT9, ACOT11) enables researchers to study loss-of-function phenotypes in cell lines and animal models. For instance, liver-specific ACOT9 knockout mice exhibit altered hepatic glucose production and lipid metabolism. Knockout models are essential for determining the physiological necessity of these enzymes.
Point Mutation
Point mutations can be introduced to mimic or abolish post-translational modifications. For example, the ACSS1-K635 acetylation knock-in mouse was generated by CRISPR to study the effect of acetylation on liver lipid metabolism during a ketogenic diet. Such models provide mechanistic insights into how specific residues regulate enzyme activity.
Knock-in
Knock-in of tagged or reporter alleles allows for real-time tracking of enzyme expression and localization. A tagged ACOT9 knock-in could be used to study its mitochondrial trafficking and interaction partners. Knock-in models are also valuable for humanizing target genes in mice for drug testing.
Overexpression
CRISPR activation (CRISPRa) or transgenic overexpression of short-chain acyl-CoA hydrolases can reveal gain-of-function effects on lipid metabolism. Overexpression of ACOT9 in hepatocytes promotes de novo lipogenesis and glucose production. Overexpression in microbial systems has been used to enhance short-chain alkane production.
How EDITGENE Supports short-chain fatty acyl-CoA hydrolase activity Research
Researchers studying short-chain fatty acyl-CoA hydrolase activity-related genes often need to determine whether a candidate gene is causally involved in metabolic phenotypes. EDITGENE provides comprehensive CRISPR-based services to generate knockout, point-mutation, knock-in, and overexpression models, as well as library screening and bioinformatics support, enabling rigorous functional studies of GO:0141126.
Contact EDITGENE today to design your custom CRISPR model for short-chain fatty acyl-CoA hydrolase activity research.
Frequently Asked Questions About short-chain fatty acyl-CoA hydrolase activity
What is short-chain fatty acyl-CoA hydrolase activity?
It is an enzymatic activity (GO:0141126) that hydrolyzes short-chain fatty acyl-CoA molecules into free short-chain fatty acids, CoA, and H+, as defined by QuickGO.
What genes are involved in short-chain fatty acyl-CoA hydrolase activity?
Key genes include ACOT9, ACOT1, ACOT2, ACOT11, and PTE1p, which encode enzymes with this activity.
What is the function of ACOT9?
ACOT9 is a mitochondrial acyl-CoA thioesterase that hydrolyzes short-chain acyl-CoAs and traffics them toward de novo lipogenesis and glucose production in the liver.
How is short-chain fatty acyl-CoA hydrolase activity linked to NAFLD?
Dysregulation of this activity contributes to hepatic steatosis and insulin resistance; inhibition of RPN11 ameliorates NAFLD by affecting metabolic enzyme stability.
What diseases are associated with short-chain fatty acyl-CoA hydrolase dysfunction?
Nonalcoholic fatty liver disease, insulin resistance, and metabolic disorders are associated with altered short-chain acyl-CoA hydrolysis.
How can CRISPR be used to study short-chain fatty acyl-CoA hydrolase activity?
CRISPR knockout, point mutation, knock-in, and overexpression models allow researchers to dissect the physiological roles of enzymes like ACOT9 and ACSS1.
What is the substrate specificity of short-chain fatty acyl-CoA hydrolases?
They preferentially hydrolyze acyl-CoAs with aliphatic tails of fewer than 6 carbons, distinguishing them from medium- and long-chain hydrolases.
Where are short-chain fatty acyl-CoA hydrolases localized?
They are found in mitochondria, peroxisomes, and brown adipose tissue, depending on the specific enzyme.
What is the role of PTE1p in yeast?
PTE1p is a peroxisomal acyl-CoA thioesterase required for efficient degradation of short straight and branched chain fatty acids in Saccharomyces cerevisiae.
Can short-chain fatty acyl-CoA hydrolases be targeted for biofuel production?
Yes, microbial production of short-chain alkanes involves acyl-CoA hydrolysis, and engineering these enzymes can enhance biofuel yields.
Conclusion
Short-chain fatty acyl-CoA hydrolase activity (GO:0141126) is a fundamental enzymatic function that regulates short-chain acyl-CoA pools and impacts diverse metabolic pathways, from hepatic lipogenesis to thermogenesis and microbial alkane production. Dysregulation of this activity is linked to NAFLD, insulin resistance, and other metabolic disorders, making it a promising therapeutic target. CRISPR-based models, including knockout, point mutation, knock-in, and overexpression, are indispensable for dissecting the precise roles of these enzymes in health and disease. EDITGENE offers comprehensive services to accelerate such research, from custom cell line generation to high-throughput screening and bioinformatics analysis.
References
- 1. Zhou B et al.. 2024. Amelioration of nonalcoholic fatty liver disease by inhibiting the deubiquitylating enzyme RPN11.. Cell Metab 36(10):2228-2244.e7 PMID: 39146936
- 2. Xu G et al.. 2025. Mitochondrial ACSS1-K635 acetylation knock-in mice exhibit altered liver lipid metabolism on a ketogenic diet.. Free Radic Biol Med 232:260-268 PMID: 40074187
- 4. Steensels S et al.. 2020. Acyl-Coenzyme A Thioesterase 9 Traffics Mitochondrial Short-Chain Fatty Acids Toward De Novo Lipogenesis and Glucose Production in the Liver.. Hepatology 72(3):857-872 PMID: 32498134
- 5. Maeda I et al.. 2006. The peroxisomal Acyl-CoA thioesterase Pte1p from Saccharomyces cerevisiae is required for efficient degradation of short straight chain and branched chain fatty acids.. J Biol Chem 281(17):11729-35 PMID: 16490786
- 6. Svensson LT et al.. 1996. Characterization and isolation of enzymes that hydrolyze short-chain acyl-CoA in rat-liver mitochondria.. Eur J Biochem 239(2):526-31 PMID: 8706763
- 7. Alexson SE et al.. 1988. A novel type of short- and medium-chain acyl-CoA hydrolases in brown adipose tissue mitochondria.. J Biol Chem 263(27):13564-71 PMID: 2901416
- 8. Choi YJ et al.. 2013. Microbial production of short-chain alkanes.. Nature 502(7472):571-4 PMID: 24077097
