GO:0004019 adenylosuccinate synthase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0004019 adenylosuccinate synthase activity is a molecular function that catalyzes the first committed step of de novo AMP biosynthesis, converting IMP and aspartate to adenylosuccinate.
• The enzyme is a validated drug target in pathogens such as Helicobacter pylori and Mycobacterium tuberculosis, and its inhibition blocks purine salvage and growth.
• In humans, ADSS1 and ADSS2 are the two adenylosuccinate synthase paralogs; ADSS2 loss re-sensitizes acute myeloid leukemia cells to BH3 mimetics.
• ADSS1 deficiency alters energy metabolism by promoting adipose tissue re-esterification through glycerol kinase upregulation.
• The catalytic mechanism is ordered and requires GTP, Mg2+, IMP and aspartate; the transition-state analog hadacidin is a classic inhibitor.
• CRISPR knockout, point-mutation, knock-in and overexpression models are essential to dissect isoform-specific and disease-relevant functions of adenylosuccinate synthase activity.
Description
Adenylosuccinate synthase activity (GO:0004019) is the enzymatic function that catalyzes the committed step of de novo AMP biosynthesis: the GTP-dependent condensation of inosine monophosphate (IMP) with L-aspartate to form adenylosuccinate. This reaction is the first dedicated step that channels purine intermediates toward adenine nucleotides, and it is therefore a central node in cellular energy and nucleic acid metabolism. In pathogens, the same activity is essential for purine salvage and growth, making it a target for antimicrobial development. In humans, two genes, ADSS1 and ADSS2, encode adenylosuccinate synthase paralogs with distinct tissue distributions and physiological roles. Recent work has linked ADSS1 to adipose energy metabolism and ADSS2 to chemotherapy sensitivity in acute myeloid leukemia, underscoring the biomedical importance of this GO term. Researchers studying GO:0004019 need robust genetic models to determine whether the enzyme is causally involved in these phenotypes, and CRISPR-based approaches are now the standard for such causal tests.
adenylosuccinate synthase activity At A Glance
| GO ID | GO:0004019 |
|---|---|
| GO term | adenylosuccinate synthase activity |
| Ontology | molecular_function |
| Synonym | IMP:L-aspartate ligase (GDP-forming) activity; adenylosuccinate synthetase activity |
| Major function | Catalyzes the GTP-dependent conversion of IMP and L-aspartate to adenylosuccinate, the first committed step of de novo AMP biosynthesis |
| Cofactor | Mg2+ and GTP are required for catalysis |
| Inhibitors | Hadacidin and its structural analogs inhibit the enzyme |
| Human genes | ADSS1 and ADSS2 encode the two human adenylosuccinate synthase paralogs |
| Pathogen relevance | Essential for growth and purine salvage in Helicobacter pylori, Mycobacterium tuberculosis and Plasmodium falciparum |
What Is GO:0004019?
Adenylosuccinate synthase activity (GO:0004019) is defined as the catalysis of the reaction IMP + L-aspartate + GTP = adenylosuccinate + GDP + phosphate. In other words, it is the molecular function that joins aspartate to IMP to build adenylosuccinate, the immediate precursor of AMP, using GTP as an energy source. This activity is the first committed and rate-limiting step of the de novo AMP branch and is also used in purine salvage in some organisms.
Why Is adenylosuccinate synthase activity Important in Cell Biology?
Adenylosuccinate synthase activity (GO:0004019) is important because it controls the first committed step of de novo AMP biosynthesis, a pathway that supplies adenine nucleotides for DNA, RNA, ATP and signaling. Because the reaction is essential in many pathogens but has distinct isoforms in humans, it is an attractive target for selective antimicrobial and antiparasitic drugs. In human disease, ADSS2 loss sensitizes acute myeloid leukemia cells to BH3 mimetics, and ADSS1 deficiency rewires adipose energy metabolism, showing that this activity has direct therapeutic and metabolic relevance. The enzyme is also a classic model for studying transition-state analog inhibition, protein-ligand interfaces and allosteric regulation.
• Controls the first committed step of de novo AMP biosynthesis, linking purine metabolism to energy homeostasis.
• Validated drug target in Helicobacter pylori, where vitamin B6 inhibits the enzyme and bacterial growth.
• Essential purine salvage enzyme in Mycobacterium tuberculosis and Plasmodium falciparum.
• ADSS2 inhibition re-sensitizes acute myeloid leukemia to BH3 mimetics, a potential combination strategy.
• ADSS1 deficiency improves energy metabolism by promoting adipose re-esterification via glycerol kinase upregulation.
• Classic target of hadacidin, a transition-state analog used to probe enzyme mechanism.
• Provides a model system for studying protein-protein interfaces and ligand-induced conformational changes.
• Isoform-specific functions make it a paradigm for paralog dissection using CRISPR genetics.
Molecular Mechanism of adenylosuccinate synthase activity
Substrate binding and the first half-reaction
In simple terms: The enzyme first grabs IMP and GTP, then positions aspartate for attack.
Adenylosuccinate synthase binds IMP and GTP in an ordered manner, with Mg2+ coordinating the nucleotide. The enzyme then recruits L-aspartate into the active site, where the 6-oxygen of IMP is activated for nucleophilic attack by the aspartate amino group. This step is the first committed reaction of de novo AMP biosynthesis and is rate-limiting for the pathway.
Catalytic condensation and adenylosuccinate formation
In simple terms: Aspartate is stitched onto IMP to make adenylosuccinate, the direct precursor of AMP.
The catalytic condensation replaces the 6-oxo group of IMP with the aspartate amino group, yielding adenylosuccinate, GDP and inorganic phosphate. The reaction is GTP-dependent and requires Mg2+ for nucleotide positioning and transition-state stabilization. Adenylosuccinate is subsequently cleaved by adenylosuccinate lyase to produce AMP and fumarate, completing the de novo AMP branch.
Inhibition by hadacidin and transition-state analogs
In simple terms: Hadacidin mimics the reaction intermediate and jams the enzyme.
Hadacidin is a structural analog of aspartate that binds the adenylosuccinate synthase active site and inhibits catalysis. Structural modifications of hadacidin alter its inhibitory potency, providing early structure-activity relationships for the enzyme. In Dictyostelium discoideum, hadacidin inhibits growth and adenylosuccinate synthase activity, confirming target engagement in a whole organism.
Isoform-specific regulation in humans
In simple terms: Humans have two versions of the enzyme, ADSS1 and ADSS2, that do different jobs in different tissues.
ADSS1 and ADSS2 are paralogous human adenylosuccinate synthases with distinct expression patterns and physiological roles. ADSS1 deficiency promotes adipose tissue re-esterification via glycerol kinase upregulation, linking the enzyme to systemic energy metabolism. ADSS2 loss impairs de novo AMP biosynthesis and re-sensitizes acute myeloid leukemia cells to BH3 mimetics, demonstrating isoform-specific therapeutic potential.
Pathogen-specific enzyme inhibition
In simple terms: The bacterial and parasite versions of the enzyme can be blocked selectively.
Vitamin B6 inhibits Helicobacter pylori adenylosuccinate synthetase and suppresses growth of reference and antibiotic-resistant clinical strains. In Mycobacterium tuberculosis, the purine salvage pathway, including adenylosuccinate synthase, is essential and has been reviewed as a drug target. Plasmodium falciparum adenine metabolism also depends on adenylosuccinate synthase activity, making it a potential antimalarial target.
Key Genes Involved in GO:0004019 adenylosuccinate synthase activity
The following genes and proteins are directly implicated in adenylosuccinate synthase activity (GO:0004019) or its immediate pathway context.
| Gene | Major Role | Research Relevance |
|---|---|---|
| ADSS1 | Human adenylosuccinate synthase 1; catalyzes IMP + aspartate to adenylosuccinate | Deficiency alters adipose energy metabolism via glycerol kinase upregulation |
| ADSS2 | Human adenylosuccinate synthase 2; de novo AMP biosynthesis | Inhibition re-sensitizes AML to BH3 mimetics |
| ADSL | Adenylosuccinate lyase; converts adenylosuccinate to AMP and fumarate | Downstream enzyme of the same pathway |
| IMPDH1/2 | Inosine monophosphate dehydrogenase; supplies IMP for the reaction | Upstream pathway context |
| GMPS | GMP synthase; balances guanine nucleotide branch | Pathway context |
| HPRT1 | Hypoxanthine phosphoribosyltransferase; purine salvage | Pathway context |
| GART | Phosphoribosylglycinamide formyltransferase; de novo purine synthesis | Pathway context |
| ATIC | AICAR transformylase/IMP cyclohydrolase; de novo purine synthesis | Pathway context |
| PAICS | Phosphoribosylaminoimidazole carboxylase; de novo purine synthesis | Pathway context |
| PPAT | Phosphoribosyl pyrophosphate amidotransferase; first step of purine synthesis | Pathway context |
| purA | Bacterial adenylosuccinate synthase | Target in Mycobacterium tuberculosis and H. pylori |
| H. pylori purA | Adenylosuccinate synthetase in Helicobacter pylori | Inhibited by vitamin B6; growth suppression |
| P. falciparum adenylosuccinate synthase | Parasite AMP biosynthesis | Antimalarial target |
| Glycerol kinase (GK) | Upregulated in ADSS1 deficiency; promotes re-esterification | Metabolic rewiring |
| BCL-2 family proteins | Apoptosis regulators; BH3 mimetics target them | ADSS2 inhibition sensitizes AML |
| Hadacidin target | Transition-state analog inhibitor of adenylosuccinate synthase | Mechanistic probe |
| Mg2+ | Essential cofactor for catalysis | Biochemical assays |
| GTP | Energy donor and substrate | Biochemical assays |
How Is adenylosuccinate synthase activity Regulated?
Adenylosuccinate synthase activity is regulated at multiple levels. The reaction requires GTP and Mg2+, and the enzyme follows an ordered kinetic mechanism in which IMP and GTP bind before aspartate. In pathogens, the enzyme is inhibited by transition-state analogs such as hadacidin, and vitamin B6 inhibits Helicobacter pylori adenylosuccinate synthetase, linking cofactor availability to activity. In humans, the two paralogs ADSS1 and ADSS2 are differentially expressed and regulated, with ADSS1 deficiency altering glycerol kinase expression and adipose re-esterification, and ADSS2 loss impairing de novo AMP biosynthesis in leukemia cells. These observations indicate that adenylosuccinate synthase activity is controlled by substrate availability, cofactors, isoform-specific expression and pharmacological inhibition.
adenylosuccinate synthase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| ADSS2 | Acute myeloid leukemia; BH3 mimetic resistance | ADSS2 knockout AML cell lines and xenografts |
| ADSS1 | Metabolic disorder; adipose energy metabolism | Adss1 knockout mouse and adipocyte models |
| H. pylori purA | Antibiotic-resistant H. pylori infection | purA knockout and vitamin B6 inhibition assays |
| M. tuberculosis purA | Tuberculosis; purine salvage dependency | purA knockout and inhibitor screens |
| P. falciparum adenylosuccinate synthase | Malaria; adenine metabolism | Parasite knockout and antimalarial assays |
Acute myeloid leukemia and BH3 mimetic sensitivity
Inhibition of ADSS2-mediated de novo AMP biosynthesis re-sensitizes acute myeloid leukemia cells to BH3 mimetics, suggesting that targeting adenylosuccinate synthase activity can overcome apoptosis resistance. This links GO:0004019 directly to cancer therapy and provides a rationale for combination strategies.
Metabolic disorders and ADSS1 deficiency
ADSS1 deficiency improves energy metabolism by promoting adipose tissue re-esterification via glycerol kinase upregulation, indicating that loss of adenylosuccinate synthase activity can remodel systemic lipid handling. This connects GO:0004019 to metabolic disease and energy homeostasis.
Bacterial and parasitic infections
Adenylosuccinate synthase activity is essential for growth and purine salvage in Helicobacter pylori, Mycobacterium tuberculosis and Plasmodium falciparum. Vitamin B6 inhibits H. pylori adenylosuccinate synthetase and suppresses growth of antibiotic-resistant clinical strains, supporting the enzyme as an antimicrobial target.
From adenylosuccinate synthase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Is ADSS2 required for AML survival and BH3 mimetic sensitivity? | ADSS2 knockout in AML cell lines and xenografts |
| Does ADSS1 loss alter adipose energy metabolism? | Adss1 knockout mouse and primary adipocytes |
| Can point mutations in the active site abolish catalysis? | CRISPR point-mutation knock-in of catalytic residues |
| Does tagged ADSS1/ADSS2 localize to specific compartments? | Knock-in of epitope tags at endogenous loci |
| Does overexpression of ADSS2 drive de novo AMP biosynthesis? | Doxycycline-inducible overexpression in cancer cells |
| Can pathogen adenylosuccinate synthase be selectively inhibited? | Bacterial purA knockout and inhibitor assays |
How to Study the adenylosuccinate synthase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Enzyme activity assay | Adenylosuccinate formation from IMP and aspartate | Kinetics and inhibitor testing |
| CRISPR knockout | Loss-of-function phenotype | Isoform-specific causality |
| CRISPR point mutation | Catalytic residue requirement | Mechanistic dissection |
| Metabolomics | Adenylosuccinate, AMP, IMP levels | Pathway flux |
| RNA-seq | Transcriptional changes | Glycerol kinase upregulation |
| Proteomics | Protein expression and interactions | Pathway network |
| Xenograft assays | Tumor growth and drug response | BH3 mimetic combination |
| Antimicrobial susceptibility testing | Bacterial growth inhibition | H. pylori inhibitor testing |
Biochemical enzyme assays
Adenylosuccinate synthase activity is measured by coupling the formation of adenylosuccinate to absorbance or fluorescence readouts, using IMP, aspartate, GTP and Mg2+. These assays are used to determine kinetic parameters and to test inhibitors such as hadacidin and vitamin B6.
CRISPR knockout and point-mutation models
CRISPR knockout of ADSS1 or ADSS2 allows causal testing of isoform-specific functions in metabolism and cancer. Point-mutation knock-in of catalytic residues can separate enzymatic activity from scaffolding functions.
Metabolomics and flux analysis
Metabolomics and stable-isotope flux analysis measure adenylosuccinate, AMP, IMP and glycerol kinase pathway intermediates to quantify pathway activity. These methods link GO:0004019 to cellular energy and lipid metabolism.
Transcriptomics and proteomics
RNA-seq and proteomics identify downstream changes such as glycerol kinase upregulation in ADSS1 deficiency and apoptosis regulators in ADSS2-deficient leukemia cells. These approaches reveal the broader regulatory network of adenylosuccinate synthase activity.
How CRISPR Can Be Used to Study GO:0004019 adenylosuccinate synthase activity
Knockout
CRISPR knockout of ADSS1 or ADSS2 is used to test whether adenylosuccinate synthase activity is required for a phenotype, such as AML survival or adipose energy metabolism. Knockout of pathogen purA validates essentiality in H. pylori and M. tuberculosis.
Point Mutation
Point-mutation knock-in of active-site residues can abolish catalysis while preserving protein expression, allowing separation of enzymatic activity from non-catalytic functions. This is critical for assigning phenotypes specifically to GO:0004019.
Knock-in
Tagged knock-in of ADSS1 or ADSS2 at endogenous loci enables localization, interaction and stability studies under native regulation. This approach avoids artifacts from overexpression and preserves isoform-specific control.
Overexpression
Doxycycline-inducible overexpression of ADSS2 or ADSS1 tests sufficiency for de novo AMP biosynthesis and downstream phenotypes such as drug resistance or metabolic rewiring. Overexpression complements knockout data to establish causality.
How EDITGENE Supports adenylosuccinate synthase activity Research
Researchers studying adenylosuccinate synthase activity-related genes often need to determine whether a candidate gene is causally involved in a phenotype, such as leukemia drug resistance or adipose energy metabolism. EDITGENE provides publication-ready CRISPR models and bioinformatics support to answer these questions with rigor and reproducibility.
Contact EDITGENE today to design your custom CRISPR model for adenylosuccinate synthase activity research.
Related Products
| Product name | Cat.No. | Species | Gene ID | |
|---|---|---|---|---|
| ADSS2 Knockout HEK293 Cell Line | EDJ-KQ3352 | Human | 159 | Details Get a Quote |
| ADSS1 Knockout HEK293 Cell Line | EDJ-KQ8157 | Human | 122622 | Details Get a Quote |
| ADSS1 Knockout A-549 Cell Line | EDJ-KQ34069 | Human | 122622 | Details Get a Quote |
| ADSS1 Knockout HCT 116 Cell Line | EDJ-KQ34070 | Human | 122622 | Details Get a Quote |
| ADSS2 Knockout A-549 Cell Line | EDJ-KQ26346 | Human | 159 | Details Get a Quote |
| ADSS2 Knockout HCT 116 Cell Line | EDJ-KQ26348 | Human | 159 | Details Get a Quote |
| ADSS2 Knockout HeLa Cell Line | EDJ-KQ26349 | Human | 159 | Details Get a Quote |
| ADSS1 Knockout HeLa Cell Line | EDJ-KQ32730 | Human | 122622 | Details Get a Quote |
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Frequently Asked Questions About adenylosuccinate synthase activity
What is adenylosuccinate synthase activity?
Adenylosuccinate synthase activity (GO:0004019) is the molecular function that catalyzes the GTP-dependent conversion of IMP and L-aspartate to adenylosuccinate, the first committed step of de novo AMP biosynthesis.
What genes are involved in adenylosuccinate synthase activity?
The main human genes are ADSS1 and ADSS2, which encode the two adenylosuccinate synthase paralogs; pathogen orthologs include purA in bacteria and the Plasmodium falciparum enzyme.
What is the reaction catalyzed by adenylosuccinate synthase?
The enzyme catalyzes IMP + L-aspartate + GTP = adenylosuccinate + GDP + phosphate, requiring Mg2+.
Why is adenylosuccinate synthase a drug target?
It is essential for purine salvage and growth in pathogens such as Helicobacter pylori and Mycobacterium tuberculosis, and its inhibition can suppress resistant strains.
How is adenylosuccinate synthase activity inhibited?
Hadacidin and its structural analogs inhibit the enzyme, and vitamin B6 inhibits Helicobacter pylori adenylosuccinate synthetase and bacterial growth.
What is the role of ADSS2 in leukemia?
Inhibition of ADSS2-mediated de novo AMP biosynthesis re-sensitizes acute myeloid leukemia cells to BH3 mimetics.
What happens in ADSS1 deficiency?
ADSS1 deficiency improves energy metabolism by promoting adipose tissue re-esterification via glycerol kinase upregulation.
How can I study adenylosuccinate synthase activity in the lab?
Common methods include enzyme activity assays, CRISPR knockout or point-mutation models, metabolomics, RNA-seq and proteomics.
Is adenylosuccinate synthase activity present in parasites?
Yes, Plasmodium falciparum adenine metabolism depends on adenylosuccinate synthase activity, making it a potential antimalarial target.
What CRISPR models are available for adenylosuccinate synthase research?
Knockout, point-mutation, knock-in, tagged knock-in and overexpression models can be generated for ADSS1, ADSS2 and pathogen orthologs.
Conclusion
Adenylosuccinate synthase activity (GO:0004019) is a central molecular function in de novo AMP biosynthesis with direct relevance to cancer, metabolic disease and infectious disease. Its two human paralogs, ADSS1 and ADSS2, have distinct physiological roles, and pathogen orthologs are validated drug targets. CRISPR-based knockout, point-mutation, knock-in and overexpression models are essential to establish causality and to translate these findings into therapies.
References
- 1. Sun J et al.. 2026. Adenylosuccinate Synthase 1 Deficiency Improves Energy Metabolism by Promoting Adipose Tissue Re-esterification via Glycerol Kinase Upregulation.. Adv Sci (Weinh) 13(1):e06270 PMID: 41117143
- 2. He X et al.. 2026. Inhibition of ADSS2-mediated de novo AMP biosynthesis re-sensitizes acute myeloid leukemia to BH3 mimetics.. Nat Cancer 7(6):944-963 PMID: 42362999
- 3. Wojtyś MI et al.. 2024. Vitamin B6 inhibits activity of Helicobacter pylori adenylosuccinate synthetase and growth of reference and clinical, antibiotic-resistant H. pylori strains.. J Enzyme Inhib Med Chem 39(1):2372734 PMID: 39149761
- 4. Ducati RG et al.. 2011. Purine Salvage Pathway in Mycobacterium tuberculosis.. Curr Med Chem 18(9):1258-75 PMID: 21366536
- 5. Larsen TA et al.. 1998. Morphology of protein-protein interfaces.. Structure 6(4):421-7 PMID: 9562553
- 6. Mehrotra S et al.. 2010. Adenine metabolism in Plasmodium falciparum.. Exp Parasitol 125(2):147-51 PMID: 20093117
- 7. SHIGEURA HT. 1963. STRUCTURAL MODIFICATIONS OF HADACIDIN AND THEIR EFFECTS ON THE ACTIVITY OF ADENYLOSUCCINATE SYNTHETASE.. J Biol Chem 238:3999-4001 PMID: 14086736
- 8. Rossomando EF et al.. 1978. Effect of hadacidin on growth and adenylosuccinate synthetase activity of Dictyostelium discoideum.. Antimicrob Agents Chemother 14(3):476-82 PMID: 568451
