GO:0047624 adenosine-tetraphosphatase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0047624 adenosine-tetraphosphatase activity catalyzes the hydrolysis of adenosine 5'-tetraphosphate (Ap4) to ATP and phosphate.
• The reaction is a phosphohydrolase activity that removes the terminal phosphate from the tetraphosphate chain, yielding ATP and inorganic phosphate.
• In Saccharomyces cerevisiae, this activity is an inherent property of soluble exopolyphosphatase, linking it to polyphosphate metabolism.
• The Bartonella bacilliformis ialA gene encodes a Nudix hydrolase active on dinucleoside 5'-polyphosphates, providing a bacterial model for related activities.
• Adenosine-tetraphosphatase activity is implicated in nucleotide homeostasis and may influence cellular energy balance and signaling.
• Research tools include CRISPR knockout, point mutation, and overexpression models to dissect gene function in this pathway.
Description
Adenosine-tetraphosphatase activity (GO:0047624) is a molecular function defined by the catalysis of the reaction: adenosine 5'-tetraphosphate + H2O = phosphate + ATP + H+. This activity is a phosphohydrolase that specifically targets adenosine 5'-tetraphosphate (Ap4), cleaving it into ATP and inorganic phosphate. The enzyme responsible for this activity in yeast was identified as a soluble exopolyphosphatase, demonstrating that adenosine-tetraphosphatase activity is an inherent property of this enzyme. In bacteria, the ialA gene product from Bartonella bacilliformis is a Nudix hydrolase active on dinucleoside 5'-polyphosphates, suggesting that related enzymes may also exhibit adenosine-tetraphosphatase activity. Understanding this activity is important for researchers studying nucleotide metabolism, energy homeostasis, and the roles of polyphosphates in cellular processes.
adenosine-tetraphosphatase activity At A Glance
| GO ID | GO:0047624 |
|---|---|
| GO term | adenosine-tetraphosphatase activity |
| Ontology | molecular_function |
| Synonym | adenosine-tetraphosphate phosphohydrolase activity |
| Major function | Catalysis of the reaction: adenosine 5'-tetraphosphate + H2O = phosphate + ATP + H+ |
| Reaction direction | Hydrolysis of adenosine 5'-tetraphosphate |
| Substrates | Adenosine 5'-tetraphosphate (Ap4), water |
| Products | ATP, phosphate, H+ |
| Enzyme class | Phosphohydrolase (EC 3.6.1.-) |
What Is GO:0047624?
Adenosine-tetraphosphatase activity is the enzymatic catalysis of the hydrolysis of adenosine 5'-tetraphosphate (Ap4) into ATP, phosphate, and a proton. This reaction removes the terminal phosphate group from the tetraphosphate chain, converting Ap4 to ATP and inorganic phosphate. The activity is classified as a phosphohydrolase and is synonymous with adenosine-tetraphosphate phosphohydrolase activity.
Why Is adenosine-tetraphosphatase activity Important in Cell Biology?
Adenosine-tetraphosphatase activity is important because it regulates the levels of adenosine 5'-tetraphosphate, a nucleotide that can influence energy metabolism and signaling. By converting Ap4 to ATP, this activity directly contributes to cellular ATP pools and phosphate homeostasis. In yeast, the enzyme responsible for this activity is a soluble exopolyphosphatase, linking it to polyphosphate metabolism, which is critical for stress responses and survival. In pathogenic bacteria such as Bartonella bacilliformis, a related Nudix hydrolase encoded by ialA is associated with erythrocyte invasion, suggesting that adenosine-tetraphosphatase-like activities may play roles in host-pathogen interactions. Thus, studying this activity can provide insights into fundamental nucleotide metabolism and potential therapeutic targets.
• Regulates adenosine 5'-tetraphosphate (Ap4) levels, affecting ATP and phosphate balance.
• Contributes to polyphosphate metabolism in yeast via exopolyphosphatase.
• May influence bacterial virulence through Nudix hydrolases like ialA in Bartonella bacilliformis.
• Provides a model for studying phosphohydrolase specificity and catalysis.
• Potential target for antimicrobial strategies targeting nucleotide metabolism.
• Relevant to energy homeostasis and stress responses in eukaryotic cells.
• Can be studied using CRISPR-based gene editing to dissect gene function.
• Links to broader Nudix hydrolase family functions in dinucleoside polyphosphate turnover.
Molecular Mechanism of adenosine-tetraphosphatase activity
Substrate recognition and binding
In simple terms: The enzyme grabs adenosine 5'-tetraphosphate and positions it for cleavage.
Adenosine-tetraphosphatase activity specifically recognizes adenosine 5'-tetraphosphate (Ap4) as a substrate. The enzyme binds the adenosine moiety and the tetraphosphate chain, positioning the terminal phosphate for hydrolysis. In yeast, this activity is associated with soluble exopolyphosphatase, which can accommodate various polyphosphate substrates, but the specific recognition of Ap4 leads to ATP production.
Catalytic hydrolysis
In simple terms: Water attacks the phosphate chain, breaking it to release ATP and phosphate.
The catalytic mechanism involves the nucleophilic attack of water on the terminal phosphate of Ap4, resulting in the cleavage of the phosphoanhydride bond and release of ATP and inorganic phosphate. This reaction is characteristic of phosphohydrolases, which use metal ions or other cofactors to activate water. The exact catalytic residues are not fully defined for all enzymes, but the reaction is known to produce ATP and phosphate as products.
Enzyme classes and families
In simple terms: Different enzymes can perform this activity, including exopolyphosphatases and Nudix hydrolases.
Adenosine-tetraphosphatase activity is not restricted to a single enzyme family. In Saccharomyces cerevisiae, it is an inherent property of soluble exopolyphosphatase, an enzyme primarily known for degrading polyphosphate. In Bartonella bacilliformis, the ialA gene encodes a Nudix hydrolase active on dinucleoside 5'-polyphosphates, which may also exhibit adenosine-tetraphosphatase activity. This suggests that multiple protein scaffolds can support this catalytic function.
Regulation and cellular context
In simple terms: The activity can be regulated by cellular conditions and may affect energy balance.
The regulation of adenosine-tetraphosphatase activity is not well characterized, but it is likely influenced by substrate availability and cellular energy status. In yeast, exopolyphosphatase activity is regulated in response to phosphate availability and stress. In bacteria, Nudix hydrolases like ialA are often regulated during infection, suggesting a role in adaptation to host environments. Further studies are needed to elucidate specific regulatory mechanisms.
Key Genes Involved in GO:0047624 adenosine-tetraphosphatase activity
The following genes and proteins are associated with adenosine-tetraphosphatase activity or related phosphohydrolase functions.
| Gene | Major Role | Research Relevance |
|---|---|---|
| PPX1 (S. cerevisiae) | Soluble exopolyphosphatase with adenosine-tetraphosphatase activity | Model for studying polyphosphate and Ap4 metabolism |
| ialA (Bartonella bacilliformis) | Nudix hydrolase active on dinucleoside 5'-polyphosphates | Linked to erythrocyte invasion and virulence |
| NUDT family (human) | Nudix hydrolases that hydrolyze dinucleoside polyphosphates | Potential orthologs with similar activities |
| ATP synthase | Produces ATP, product of the reaction | Energy metabolism context |
| Inorganic pyrophosphatase | Hydrolyzes pyrophosphate, related to phosphate metabolism | May interact with Ap4 pathways |
| Polyphosphate kinase | Synthesizes polyphosphate, related to exopolyphosphatase | Opposing enzyme in polyP metabolism |
| Adenylate kinase | Interconverts adenine nucleotides | Maintains nucleotide balance |
| Nudix hydrolase family | Broad family of phosphohydrolases | Includes enzymes acting on Ap4 and related substrates |
| Exopolyphosphatase (other organisms) | Degrades polyphosphate | Potential adenosine-tetraphosphatase activity |
| Ap4A hydrolase | Hydrolyzes diadenosine tetraphosphate | Related dinucleoside polyphosphate metabolism |
| ATP-binding cassette transporters | Transport nucleotides | May influence substrate availability |
| Phosphate transporters | Uptake phosphate | Regulate phosphate homeostasis |
| Protein kinases | Phosphorylate proteins using ATP | Downstream of ATP produced |
| Adenosine deaminase | Metabolizes adenosine | Affects adenosine pools |
| 5'-Nucleotidase | Produces adenosine from AMP | Related to adenosine metabolism |
| Nudix motif proteins | Contain Nudix fold for phosphohydrolase activity | Candidate adenosine-tetraphosphatases |
How Is adenosine-tetraphosphatase activity Regulated?
The regulation of adenosine-tetraphosphatase activity is not extensively studied. In yeast, the soluble exopolyphosphatase responsible for this activity is regulated by phosphate availability and cellular stress responses. In Bartonella bacilliformis, the expression of ialA, which encodes a Nudix hydrolase, may be controlled during infection, but specific regulatory pathways remain to be defined. General mechanisms such as substrate availability, post-translational modifications, and metal ion cofactors likely influence activity, but further research is needed.
adenosine-tetraphosphatase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| ialA | Bartonella bacilliformis infection | Knockout in Bartonella, infection assays |
| PPX1 | Phosphate metabolism disorders | Yeast knockout and overexpression |
| NUDT family | Cancer, metabolic disorders | Human cell line knockout |
| Exopolyphosphatase | Polyphosphate-related diseases | Mouse models with gene deletion |
| Nudix hydrolases | Inflammatory and immune disorders | CRISPR knock-in of mutations |
Bacterial infection and virulence
The ialA gene from Bartonella bacilliformis, which encodes a Nudix hydrolase active on dinucleoside 5'-polyphosphates, is associated with the invasion of human erythrocytes. This suggests that adenosine-tetraphosphatase-like activities may contribute to bacterial pathogenesis. Targeting such enzymes could provide new strategies for treating infections caused by Bartonella species.
Metabolic disorders and phosphate imbalance
Adenosine-tetraphosphatase activity produces ATP and phosphate, directly impacting cellular energy and phosphate homeostasis. Dysregulation of this activity could contribute to metabolic disorders characterized by altered phosphate levels or ATP imbalance. However, direct links to human disease remain to be established, and most evidence comes from yeast models.
Cancer and nucleotide metabolism
Altered nucleotide metabolism is a hallmark of cancer, and enzymes that regulate ATP and phosphate levels may influence tumor growth. While adenosine-tetraphosphatase activity has not been directly linked to cancer, related Nudix hydrolases are implicated in various cancers. Further research is needed to determine if this activity plays a role in oncogenesis.
From adenosine-tetraphosphatase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does PPX1 knockout abolish adenosine-tetraphosphatase activity? | Yeast knockout (CRISPR or homologous recombination) |
| What is the effect of ialA point mutations on enzyme activity? | Bacterial point mutation via CRISPR |
| Can human NUDT genes complement yeast ppx1 deletion? | Knock-in of human NUDT into yeast |
| How does overexpression of exopolyphosphatase affect Ap4 levels? | Overexpression in yeast or mammalian cells |
| What is the subcellular localization of adenosine-tetraphosphatase? | Tagged knock-in with fluorescent protein |
| Does adenosine-tetraphosphatase activity influence infection? | Bacterial knockout in infection models |
How to Study the adenosine-tetraphosphatase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Malachite green assay | Inorganic phosphate release | Enzyme kinetics |
| Luciferase assay | ATP production | Activity detection |
| CRISPR knockout | Gene function | Loss-of-function studies |
| Overexpression | Enzyme levels and activity | Gain-of-function studies |
| Mass spectrometry | Ap4 and ATP quantification | Metabolite profiling |
| Western blot | Protein expression | Validation of knockout/overexpression |
| Fluorescence microscopy | Subcellular localization | Tagged knock-in |
| Infection assays | Bacterial virulence | ialA function |
Enzymatic assays
Adenosine-tetraphosphatase activity can be measured using coupled enzymatic assays that detect phosphate release or ATP formation. For example, the release of inorganic phosphate from Ap4 can be quantified using malachite green or molybdate-based assays. ATP production can be monitored using luciferase-based luminescence.
Genetic knockout and knockdown
CRISPR-Cas9 knockout of candidate genes such as PPX1 in yeast or ialA in bacteria can abolish adenosine-tetraphosphatase activity, allowing researchers to confirm gene function. Knockdown using RNA interference can also be used in mammalian cells to reduce expression and assess activity.
Overexpression and purification
Recombinant enzymes can be overexpressed in E. coli or yeast and purified for in vitro assays. This allows detailed kinetic characterization of adenosine-tetraphosphatase activity, including Km and Vmax determination. Overexpression in cells can also elevate activity and alter nucleotide pools.
Metabolite profiling
Mass spectrometry-based metabolomics can quantify adenosine 5'-tetraphosphate and ATP levels in cells with altered enzyme expression. This provides insights into the physiological role of the activity. Such profiling can be combined with isotope labeling to trace flux through the pathway.
How CRISPR Can Be Used to Study GO:0047624 adenosine-tetraphosphatase activity
Knockout
CRISPR-Cas9 knockout of PPX1 in Saccharomyces cerevisiae or ialA in Bartonella bacilliformis can eliminate adenosine-tetraphosphatase activity, enabling researchers to study its physiological consequences. Knockout cell lines can be used to assess changes in Ap4 and ATP levels, as well as effects on growth and stress responses.
Point Mutation
Introducing point mutations in catalytic residues of candidate enzymes via CRISPR base editing or homology-directed repair can dissect the mechanism of adenosine-tetraphosphatase activity. For example, mutating the Nudix motif in ialA can abolish hydrolase activity and test its role in erythrocyte invasion.
Knock-in
Knock-in of tagged versions of PPX1 or ialA using CRISPR can facilitate localization and interaction studies. Fluorescent tags allow live-cell imaging of the enzyme, while affinity tags enable purification for biochemical assays. Knock-in of human NUDT genes into yeast can test functional conservation.
Overexpression
CRISPR activation (CRISPRa) or plasmid-based overexpression can increase adenosine-tetraphosphatase activity in cells. This is useful for studying the effects of elevated Ap4 hydrolysis on nucleotide pools and cellular metabolism. Overexpression in bacterial pathogens can also enhance virulence in infection models.
How EDITGENE Supports adenosine-tetraphosphatase activity Research
Researchers studying adenosine-tetraphosphatase activity-related genes often need to determine whether a candidate gene is causally involved in the hydrolysis of adenosine 5'-tetraphosphate and how its loss or gain of function affects cellular phenotypes. EDITGENE provides a comprehensive suite of CRISPR-based services to accelerate this research, from gene knockout to precise point mutations and knock-in models.
Contact EDITGENE today to design your custom CRISPR model for adenosine-tetraphosphatase activity research.
Frequently Asked Questions About adenosine-tetraphosphatase activity
What is adenosine-tetraphosphatase activity?
Adenosine-tetraphosphatase activity (GO:0047624) is the enzymatic catalysis of the reaction: adenosine 5'-tetraphosphate + H2O = phosphate + ATP + H+, effectively hydrolyzing Ap4 to ATP and phosphate.
What genes are involved in adenosine-tetraphosphatase activity?
Genes include PPX1 in Saccharomyces cerevisiae, which encodes a soluble exopolyphosphatase with this activity, and ialA in Bartonella bacilliformis, which encodes a Nudix hydrolase active on dinucleoside polyphosphates.
What is the reaction catalyzed by adenosine-tetraphosphatase?
The enzyme catalyzes the hydrolysis of adenosine 5'-tetraphosphate to ATP, phosphate, and a proton.
Which enzyme classes exhibit adenosine-tetraphosphatase activity?
Both exopolyphosphatases and Nudix hydrolases can exhibit this activity, as shown for yeast exopolyphosphatase and bacterial Nudix hydrolase ialA.
How is adenosine-tetraphosphatase activity measured?
It can be measured by detecting phosphate release using malachite green or by monitoring ATP production with luciferase assays.
Is adenosine-tetraphosphatase activity linked to disease?
The bacterial ialA gene is associated with erythrocyte invasion, suggesting a role in infection, but direct links to human diseases are not well established.
What is the role of adenosine-tetraphosphatase in yeast?
In yeast, it is an inherent property of soluble exopolyphosphatase, contributing to polyphosphate metabolism and phosphate homeostasis.
Can CRISPR be used to study adenosine-tetraphosphatase activity?
Yes, CRISPR knockout, point mutation, and knock-in models can be used to dissect gene function and enzymatic activity.
What are the substrates and products of adenosine-tetraphosphatase?
The substrate is adenosine 5'-tetraphosphate and water; the products are ATP, phosphate, and H+.
Where can I find validated CRISPR models for adenosine-tetraphosphatase research?
EDITGENE provides custom CRISPR services including knockout, point mutation, knock-in, overexpression, and library screening for genes related to this activity.
Conclusion
Adenosine-tetraphosphatase activity (GO:0047624) is a specific phosphohydrolase function that converts adenosine 5'-tetraphosphate to ATP and phosphate. It is carried out by enzymes such as yeast exopolyphosphatase and bacterial Nudix hydrolases, linking it to polyphosphate metabolism and infection biology. Understanding this activity can provide insights into nucleotide homeostasis and potential therapeutic targets. EDITGENE offers comprehensive CRISPR solutions to study the genes responsible for this activity, from knockout to precise knock-in models.
References
- 1. Guranowski A et al.. 1998. Adenosine 5'-tetraphosphate phosphohydrolase activity is an inherent property of soluble exopolyphosphatase from yeast Saccharomyces cerevisiae.. Biochim Biophys Acta 1380(2):232-8 PMID: 9565693
- 2. Conyers GB et al.. 1999. The gene, ialA, associated with the invasion of human erythrocytes by Bartonella bacilliformis, designates a nudix hydrolase active on dinucleoside 5'-polyphosphates.. J Biol Chem 274(3):1203-6 PMID: 9880487
