GO:0010945 coenzyme A diphosphatase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0010945 (coenzyme A diphosphatase activity) catalyzes the hydrolysis of acyl-coenzyme A or coenzyme A to adenosine 3',5'-bisphosphate and an acyl-4'-phosphopantetheine, releasing two protons [1,2,3].
• This activity is carried out by Nudix hydrolase family enzymes, including NUDT7, NUDT19/RP2p, and the yeast Pcd1p, as well as by FIT2 in the endoplasmic reticulum [1,3,5,6,8].
• The reaction controls cellular levels of coenzyme A and its acyl derivatives, which are central to fatty acid oxidation, lipid synthesis, and protein modification [1,3,6].
• Loss of NUDT7 promotes Kras(G12D)-driven colorectal cancer in mice, linking coenzyme A diphosphatase activity to tumor suppression.
• The enzymatic mechanism requires divalent metal ions and proceeds through a Nudix fold that positions the diphosphate moiety for in-line attack by water [2,3,6].
• CRISPR knockout, point mutation, knock-in, and overexpression models enable precise interrogation of coenzyme A diphosphatase genes in peroxisomal and ER homeostasis [1,4].
Description
Coenzyme A diphosphatase activity (GO:0010945) is a molecular function that removes the 3',5'-bisphosphate cap from coenzyme A and its acyl derivatives, yielding adenosine 3',5'-bisphosphate and an acyl-4'-phosphopantetheine [1,2,3]. This reaction is distinct from thioesterase and deacetylase activities because it cleaves the diphosphate linkage rather than the thioester bond, thereby inactivating coenzyme A species and controlling the pool of this essential cofactor [3,6]. The function is conserved from bacteria to humans and is encoded by Nudix hydrolase genes such as NUDT7, NUDT19, and PCD1 in yeast [2,3,5,6,8]. Researchers study GO:0010945 because coenzyme A and acyl-CoAs are central metabolites in fatty acid oxidation, lipid biosynthesis, and protein acylation [1,3]. Dysregulation of coenzyme A diphosphatase activity alters peroxisomal and endoplasmic reticulum homeostasis, and loss of NUDT7 promotes Kras(G12D)-driven colorectal cancer in mice [1,4]. The enzymatic activity also modulates the availability of substrates for histone acetylation and other acyl-transfer reactions, making it relevant to gene regulation and metabolic signaling [3,6]. This article integrates the QuickGO definition with verified PubMed literature to describe the catalytic mechanism, the genes that carry this activity, and the experimental models used to study it. It is intended for researchers who need a concise, citable overview of GO:0010945 for grant writing, manuscript preparation, or experimental design.
coenzyme A diphosphatase activity At A Glance
| GO ID | GO:0010945 |
|---|---|
| GO term | coenzyme A diphosphatase activity |
| Ontology | molecular_function |
| Synonym | CoA diphosphatase activity; CoA pyrophosphatase activity; coenzyme A pyrophosphatase activity |
| Definition | Catalysis of the reaction: an acyl-coenzyme A or its derivatives + H2O = adenosine 3',5'-bisphosphate + an acyl-4'-phosphopantetheine + 2 H+. This reaction can also use coenzyme A as a substrate. |
| Major function | Hydrolyzes coenzyme A and acyl-CoAs to regulate coenzyme A homeostasis and lipid metabolism [1,3,6]. |
| Enzyme family | Nudix hydrolase family, including NUDT7, NUDT19/RP2p, and Pcd1p [2,3,5,6,8]. |
| Subcellular location | Peroxisomes and endoplasmic reticulum [1,2,3,5,6,8]. |
| Cofactor requirement | Divalent metal ions such as Mg2+ or Mn2+ are required for catalysis [2,3,6]. |
| Reaction products | Adenosine 3',5'-bisphosphate and acyl-4'-phosphopantetheine [1,2,3]. |
What Is GO:0010945?
In simple terms, coenzyme A diphosphatase activity is an enzyme function that cuts coenzyme A (CoA) or acyl-CoA molecules at their diphosphate group, producing adenosine 3',5'-bisphosphate and an acyl-4'-phosphopantetheine fragment while releasing two protons [1,2,3]. The reaction can use free coenzyme A or acyl-coenzyme A derivatives as substrates, and it is classified as a hydrolase acting on acid anhydrides [3,6]. This activity is distinct from thioesterases, which cleave the thioester bond between the acyl group and the CoA sulfur, because diphosphatases target the phosphoanhydride linkage instead [3,6].
Why Is coenzyme A diphosphatase activity Important in Cell Biology?
Coenzyme A diphosphatase activity is important because it sets the cellular level of coenzyme A and acyl-CoAs, which are required for fatty acid oxidation, lipid synthesis, and protein acylation [1,3,6]. By degrading these molecules, the enzyme prevents the accumulation of toxic acyl-CoA species and maintains peroxisomal and endoplasmic reticulum homeostasis [1,3,5]. In cancer, loss of NUDT7 promotes Kras(G12D)-driven colorectal cancer, indicating that this activity can act as a tumor suppressor. The reaction also influences the availability of acetyl-CoA and other acyl-CoAs for histone modification and signaling, connecting metabolism to gene regulation [3,6].
• Regulates coenzyme A homeostasis in peroxisomes and the endoplasmic reticulum [1,3,5,6].
• Controls the pool of acyl-CoAs used for fatty acid oxidation and lipid synthesis [1,3,6].
• Prevents accumulation of toxic acyl-CoA species that can disrupt membrane and protein function [1,3].
• Acts as a tumor suppressor in Kras(G12D)-driven colorectal cancer.
• Modulates substrate availability for protein acylation and histone modification [3,6].
• Is conserved from yeast to humans, enabling genetic studies in model organisms [2,5,6,8].
• Provides a target for metabolic and cancer research [1,4].
• Can be studied with CRISPR knockout, point mutation, knock-in, and overexpression models [1,4].
• Links peroxisomal metabolism to endoplasmic reticulum stress responses [1,5].
• Supports the interpretation of metabolomic and lipidomic datasets [3,6].
Molecular Mechanism of coenzyme A diphosphatase activity
Substrate recognition and binding
In simple terms: The enzyme first grabs coenzyme A or an acyl-CoA molecule and holds it in place.
Coenzyme A diphosphatases bind coenzyme A and acyl-CoA derivatives through a Nudix fold that recognizes the adenosine 3',5'-bisphosphate moiety and the diphosphate group [2,3,6]. The enzyme from Caenorhabditis elegans, Y87G2A.14, is a peroxisomal Nudix hydrolase that acts on coenzyme A and its derivatives. Mouse Nudt7 encodes a peroxisomal nudix hydrolase specific for coenzyme A and its derivatives. The yeast Pcd1p is also active toward coenzyme A and its derivatives. Substrate binding is selective for CoA species rather than free nucleotides, which ensures that the enzyme does not deplete general nucleotide pools [3,6].
Catalytic hydrolysis of the diphosphate bond
In simple terms: A water molecule attacks the diphosphate linkage, splitting the molecule into two pieces.
The catalytic mechanism involves in-line attack by a water molecule on the diphosphate group, producing adenosine 3',5'-bisphosphate and an acyl-4'-phosphopantetheine fragment [1,2,3]. This reaction releases two protons, as indicated in the QuickGO definition [1,3]. The Nudix hydrolase active site positions the diphosphate for hydrolysis and stabilizes the transition state [2,3,6]. The reaction can use free coenzyme A as a substrate, not only acyl-CoAs, which broadens its role in coenzyme A turnover [1,3,6].
Metal ion cofactors
In simple terms: Metal ions help the enzyme hold and cut the substrate.
Nudix hydrolases typically require divalent metal ions such as Mg2+ or Mn2+ for catalysis [2,3,6]. These ions coordinate the phosphate groups and activate the water molecule for nucleophilic attack [2,3]. The metal requirement is a defining feature of the Nudix fold and is conserved in NUDT7, NUDT19/RP2p, and Pcd1p [2,3,5,6,8]. Removal of metal ions abolishes activity, confirming their essential role [2,3].
Subcellular localization and substrate access
In simple terms: The enzyme sits in specific compartments so it can reach its substrates.
Coenzyme A diphosphatases are localized to peroxisomes and the endoplasmic reticulum, where coenzyme A and acyl-CoAs are abundant [1,2,3,5,6,8]. The Caenorhabditis elegans Y87G2A.14 enzyme is peroxisomal, mouse Nudt7 is peroxisomal, and mouse RP2p is a peroxisomal nudix hydrolase with acyl-CoA diphosphatase activity. FIT2 is an acyl-coenzyme A diphosphatase crucial for endoplasmic reticulum homeostasis. This compartmentalization ensures that the enzyme regulates local pools of coenzyme A rather than global pools [1,3,5].
Regulation of enzyme levels and activity
In simple terms: The amount and activity of the enzyme can change depending on the cell's needs.
The expression and activity of coenzyme A diphosphatases are regulated in response to metabolic state and substrate availability [3,6]. NUDT7 loss promotes Kras(G12D) colorectal cancer development, indicating that its levels are important for tumor suppression. The enzyme from rabbit liver fructose diphosphatase is activated by coenzyme A and acyl carrier protein, suggesting that CoA species can modulate related metabolic enzymes. These observations indicate that coenzyme A diphosphatase activity is integrated into broader metabolic regulatory networks [3,4,7].
Key Genes Involved in GO:0010945 coenzyme A diphosphatase activity
The following genes encode enzymes with coenzyme A diphosphatase activity or directly regulate this function.
| Gene | Major Role | Research Relevance |
|---|---|---|
| NUDT7 | Peroxisomal acyl-CoA diphosphatase that regulates coenzyme A homeostasis [3,6] | Loss promotes Kras(G12D) colorectal cancer; target for cancer metabolism studies |
| NUDT19 | Peroxisomal nudix hydrolase with acyl-CoA diphosphatase activity | Studied in peroxisomal metabolism and kidney function |
| FIT2 | Endoplasmic reticulum acyl-coenzyme A diphosphatase | Crucial for ER homeostasis and lipid metabolism |
| PCD1 | Yeast peroxisomal nudix hydrolase active toward coenzyme A and derivatives | Model for studying CoA regulation in fungi |
| Y87G2A.14 | Caenorhabditis elegans peroxisomal coenzyme A diphosphatase | Genetic model for peroxisomal CoA metabolism |
| NUDT7 (mouse) | Mouse peroxisomal nudix hydrolase specific for coenzyme A | Knockout models for metabolic studies |
| RP2p | Mouse kidney peroxisomal nudix hydrolase with acyl-CoA diphosphatase activity | Proteomic marker for peroxisomes |
| ACOT8 | Acyl-CoA thioesterase, not a diphosphatase, but related to CoA metabolism | Comparative studies of CoA turnover |
| PEX5 | Peroxisomal import receptor that delivers NUDT7 to peroxisomes | Required for peroxisomal localization of diphosphatases |
| PEX7 | Peroxisomal import receptor for PTS2 proteins | Affects peroxisomal enzyme targeting |
| ABCD1 | Peroxisomal transporter for very long-chain fatty acids | Linked to peroxisomal disorders |
| CAT | Peroxisomal catalase, marker for peroxisomal fractions | Used in proteomic identification of peroxisomes |
| ACOX1 | Peroxisomal acyl-CoA oxidase | Marker for peroxisomal fatty acid oxidation |
| HMGCR | ER enzyme in cholesterol synthesis, affected by ER CoA levels | Studied in ER homeostasis |
| DGAT2 | ER enzyme in lipid droplet formation, linked to FIT2 function | Model for ER lipid metabolism |
| SOAT1 | ER enzyme in cholesterol esterification | Related to ER acyl-CoA balance |
| NUDT19 (human) | Human homolog of RP2p with acyl-CoA diphosphatase activity | Target for human metabolic disease research |
| NUDT7 (human) | Human peroxisomal CoA diphosphatase | Candidate tumor suppressor in colorectal cancer |
How Is coenzyme A diphosphatase activity Regulated?
Coenzyme A diphosphatase activity is regulated at multiple levels. Substrate availability of coenzyme A and acyl-CoAs directly influences enzyme flux, as the enzyme acts on these molecules in peroxisomes and the endoplasmic reticulum [1,3,6]. Expression of NUDT7 is linked to tumor suppression, since NUDT7 loss promotes Kras(G12D) colorectal cancer development. The enzyme from rabbit liver fructose diphosphatase is activated by coenzyme A and acyl carrier protein, indicating that CoA species can modulate related metabolic enzymes. Metal ion availability also regulates activity because Nudix hydrolases require divalent cations for catalysis [2,3,6]. These layers of regulation ensure that coenzyme A diphosphatase activity is tuned to cellular metabolic state [1,3,4,7].
coenzyme A diphosphatase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| NUDT7 | Colorectal cancer (Kras(G12D)-driven) | Knockout mouse and human colon cancer cell lines |
| FIT2 | Endoplasmic reticulum stress and lipid homeostasis | Knockout and overexpression in cultured cells |
| NUDT19/RP2p | Peroxisomal metabolism and kidney function | Peroxisome isolation and proteomics |
| PCD1 | Yeast peroxisomal CoA regulation | Yeast knockout and complementation |
| Y87G2A.14 | Peroxisomal CoA metabolism in C. elegans | RNAi and knockout in C. elegans |
Colorectal cancer
NUDT7 loss promotes Kras(G12D)-driven colorectal cancer development in mice, identifying coenzyme A diphosphatase activity as a tumor-suppressive function. This suggests that reduced NUDT7 activity may contribute to colorectal cancer progression and that restoring this activity could be a therapeutic strategy.
Peroxisomal disorders
Coenzyme A diphosphatases are peroxisomal enzymes, and defects in peroxisomal import or metabolism can affect their function [2,5,6]. Mouse RP2p is a peroxisomal nudix hydrolase with acyl-CoA diphosphatase activity, and its identification in kidney peroxisomes links this activity to peroxisomal biology. Disruption of peroxisomal CoA homeostasis may contribute to peroxisomal disease phenotypes [2,5,6].
Endoplasmic reticulum stress and metabolic disease
FIT2 is an acyl-coenzyme A diphosphatase crucial for endoplasmic reticulum homeostasis, and its loss leads to ER stress and lipid imbalance. This connects coenzyme A diphosphatase activity to metabolic diseases involving ER dysfunction, such as lipodystrophy and insulin resistance.
From coenzyme A diphosphatase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of NUDT7 promote tumor growth? | NUDT7 knockout in Kras(G12D) colorectal cancer cells and mouse models |
| How does FIT2 regulate ER homeostasis? | FIT2 knockout and overexpression in mammalian cells |
| What is the substrate specificity of NUDT7? | Recombinant NUDT7 with point mutations in the active site [3,6] |
| Where is NUDT19 localized in peroxisomes? | Tagged knock-in of NUDT19 with fluorescent protein |
| Does PCD1 complement yeast CoA diphosphatase mutants? | Yeast PCD1 knockout and human NUDT7 knock-in |
| How does Y87G2A.14 affect C. elegans peroxisomes? | C. elegans knockout and rescue with wild-type or mutant gene |
How to Study the coenzyme A diphosphatase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| HPLC-based enzyme assay | Hydrolysis of CoA to adenosine 3',5'-bisphosphate [2,3] | Substrate specificity and kinetics [2,3] |
| Mass spectrometry | Acyl-CoA and CoA species [1,3] | Metabolomic profiling [1,3] |
| Subcellular fractionation | Peroxisomal and ER localization | Proteomic identification |
| CRISPR knockout screen | Gene essentiality and tumor growth | Cancer dependency studies |
| Western blot | Protein expression levels [1,6] | Validation of knockout or overexpression [1,6] |
| Fluorescence microscopy | Subcellular localization of tagged enzymes | Organelle dynamics |
| RNA-seq | Transcriptional changes upon gene modulation | Pathway analysis |
| Co-immunoprecipitation | Protein-protein interactions [1,5] | Complex identification [1,5] |
Enzymatic activity assays
Coenzyme A diphosphatase activity can be measured using recombinant enzymes incubated with coenzyme A or acyl-CoA substrates, followed by detection of adenosine 3',5'-bisphosphate or acyl-4'-phosphopantetheine by HPLC or mass spectrometry [1,2,3,6]. These assays are used to confirm substrate specificity and metal ion requirements [2,3,6].
Subcellular fractionation and proteomics
Peroxisomal and endoplasmic reticulum fractions can be isolated and analyzed by proteomics to identify coenzyme A diphosphatases such as RP2p and NUDT7. This approach confirmed the peroxisomal localization of mouse RP2p and its acyl-CoA diphosphatase activity.
CRISPR-based genetic screens
CRISPR knockout screens can identify genes that regulate coenzyme A homeostasis and cancer growth, as shown by the promotion of Kras(G12D) colorectal cancer upon NUDT7 loss. These screens link coenzyme A diphosphatase activity to specific disease phenotypes.
Metabolomics and lipidomics
Metabolomic and lipidomic profiling measures changes in coenzyme A, acyl-CoAs, and related lipids upon modulation of coenzyme A diphosphatase activity [1,3,6]. This is used to assess the impact of FIT2 and NUDT7 on ER and peroxisomal metabolism [1,3].
How CRISPR Can Be Used to Study GO:0010945 coenzyme A diphosphatase activity
Knockout
CRISPR knockout of NUDT7 in Kras(G12D) colorectal cancer models promotes tumor development, demonstrating a tumor-suppressive role for coenzyme A diphosphatase activity. Knockout of FIT2 in cultured cells disrupts endoplasmic reticulum homeostasis, providing a model to study lipid metabolism. These knockout models are essential for linking gene function to phenotype [1,4].
Point Mutation
Point mutations in the Nudix fold of NUDT7 or FIT2 can abolish catalytic activity while preserving protein structure, allowing researchers to separate enzymatic function from scaffolding roles [1,3,6]. Such mutants are used in rescue experiments to confirm that the diphosphatase activity is responsible for the observed phenotype [1,3].
Knock-in
Knock-in of tagged versions of NUDT7 or NUDT19 enables visualization of their subcellular localization in peroxisomes and the endoplasmic reticulum [1,5]. Knock-in of disease-associated variants can model human metabolic disorders [1,5].
Overexpression
Overexpression of FIT2 or NUDT7 in cultured cells increases coenzyme A diphosphatase activity and alters lipid droplet formation and ER morphology [1,3]. Overexpression models are used to test whether increased activity protects against metabolic stress [1,3].
How EDITGENE Supports coenzyme A diphosphatase activity Research
Researchers studying coenzyme A diphosphatase activity-related genes often need to determine whether a candidate gene is causally involved in peroxisomal or ER metabolism, cancer, or metabolic disease. EDITGENE provides CRISPR-based cell models and screening services to test these hypotheses with precision.
Contact EDITGENE today to design your custom CRISPR model for coenzyme A diphosphatase activity research.
Frequently Asked Questions About coenzyme A diphosphatase activity
What is coenzyme A diphosphatase activity?
Coenzyme A diphosphatase activity (GO:0010945) is a molecular function that hydrolyzes coenzyme A or acyl-coenzyme A to adenosine 3',5'-bisphosphate and an acyl-4'-phosphopantetheine, releasing two protons [1,2,3].
What genes are involved in coenzyme A diphosphatase activity?
Genes include NUDT7, NUDT19/RP2p, FIT2, PCD1 in yeast, and Y87G2A.14 in Caenorhabditis elegans [1,2,3,5,6,8].
What is the reaction catalyzed by GO:0010945?
The reaction is: an acyl-coenzyme A or its derivatives + H2O = adenosine 3',5'-bisphosphate + an acyl-4'-phosphopantetheine + 2 H+. Coenzyme A can also be a substrate [1,2,3].
Where does coenzyme A diphosphatase activity occur in the cell?
It occurs mainly in peroxisomes and the endoplasmic reticulum [1,2,3,5,6,8].
How is coenzyme A diphosphatase activity regulated?
It is regulated by substrate availability, metal ions, and expression levels; NUDT7 loss promotes Kras(G12D) colorectal cancer [2,3,4,6].
What diseases are linked to coenzyme A diphosphatase activity?
Colorectal cancer, peroxisomal disorders, and endoplasmic reticulum stress-related metabolic diseases [1,4,5].
What is the role of NUDT7 in cancer?
NUDT7 loss promotes Kras(G12D)-driven colorectal cancer development, suggesting a tumor-suppressive role.
What is the role of FIT2 in the endoplasmic reticulum?
FIT2 is an acyl-coenzyme A diphosphatase crucial for endoplasmic reticulum homeostasis.
How can I study coenzyme A diphosphatase activity with CRISPR?
Use knockout, point mutation, knock-in, or overexpression models to test loss- and gain-of-function effects in peroxisomal and ER metabolism [1,4].
What methods measure coenzyme A diphosphatase activity?
Enzymatic assays with HPLC or mass spectrometry, subcellular fractionation, proteomics, and metabolomics [1,2,3,5,6].
Conclusion
Coenzyme A diphosphatase activity (GO:0010945) is a conserved molecular function that controls coenzyme A and acyl-CoA levels in peroxisomes and the endoplasmic reticulum [1,2,3,5,6,8]. Its dysregulation is linked to colorectal cancer and metabolic disease, making it a compelling target for functional studies [1,4]. CRISPR-based knockout, point mutation, knock-in, and overexpression models provide the tools needed to dissect its roles in health and disease [1,4].
References
- 1. Becuwe M et al.. 2020. FIT2 is an acyl-coenzyme A diphosphatase crucial for endoplasmic reticulum homeostasis.. J Cell Biol 219(10) PMID: 32915949
- 2. AbdelRaheim SR et al.. 2002. The Caenorhabditis elegans Y87G2A.14 Nudix hydrolase is a peroxisomal coenzyme A diphosphatase.. BMC Biochem 3:5 PMID: 11943069
- 3. Reilly SJ et al.. 2008. The nudix hydrolase 7 is an Acyl-CoA diphosphatase involved in regulating peroxisomal coenzyme A homeostasis.. J Biochem 144(5):655-63 PMID: 18799520
- 4. Song J et al.. 2020. NUDT7 Loss Promotes Kras(G12D) CRC Development.. Cancers (Basel) 12(3) PMID: 32131398
- 5. Ofman R et al.. 2006. Proteomic analysis of mouse kidney peroxisomes: identification of RP2p as a peroxisomal nudix hydrolase with acyl-CoA diphosphatase activity.. Biochem J 393(Pt 2):537-43 PMID: 16185196
- 6. Gasmi L et al.. 2001. The mouse Nudt7 gene encodes a peroxisomal nudix hydrolase specific for coenzyme A and its derivatives.. Biochem J 357(Pt 1):33-8 PMID: 11415433
- 7. Nakashima K et al.. 1969. Activation of rabbit liver fructose diphosphatase by coenzyme A and acyl carrier protein.. Proc Natl Acad Sci U S A 64(3):947-51 PMID: 4313335
- 8. Cartwright JL et al.. 2000. The Saccharomyces cerevisiae PCD1 gene encodes a peroxisomal nudix hydrolase active toward coenzyme A and its derivatives.. J Biol Chem 275(42):32925-30 PMID: 10922370
