GO:0047777 (S)-citramalyl-CoA lyase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0047777 describes the enzymatic activity that cleaves (3S)-citramalyl-CoA into acetyl-CoA and pyruvate, a central step in bacterial itaconate degradation and in the autotrophic 3-hydroxypropionate cycle.
• The reaction is a carbon-carbon bond cleavage (lyase) that funnels itaconate-derived carbon into central metabolism, supporting pathogenicity of Mycobacterium tuberculosis and other bacteria.
• The enzyme belongs to the itaconate degradation gene cluster in bacteria such as Yersinia pestis, Pseudomonas species, and Burkholderia xenovorans.
• In Chloroflexus aurantiacus, the enzyme (R-citramalyl-CoA lyase) participates in the autotrophic 3-hydroxypropionate cycle for CO2 fixation.
• Loss of (S)-citramalyl-CoA lyase activity impairs itaconate dissimilation and reduces bacterial survival in macrophages, linking the activity to host-pathogen interactions.
• CRISPR knockout, point-mutation, knock-in, and overexpression models enable precise dissection of this activity in microbial and host-cell contexts.
Description
GO:0047777, (S)-citramalyl-CoA lyase activity, is a molecular function defined by the catalysis of the reaction (3S)-citramalyl-CoA = acetyl-CoA + pyruvate [QuickGO]. This activity is a key enzymatic step in bacterial itaconate degradation, a pathway that allows pathogens such as Mycobacterium tuberculosis and Yersinia pestis to metabolize the host-derived antimicrobial metabolite itaconate. The enzyme is also part of the autotrophic 3-hydroxypropionate cycle in Chloroflexus aurantiacus, where it contributes to carbon fixation. Understanding this activity is important because it connects central carbon metabolism, bacterial virulence, and potential biotechnological applications in itaconate production and degradation. Researchers studying this term often need to determine whether a candidate gene encodes a functional (S)-citramalyl-CoA lyase, how its activity is regulated, and how it influences microbial physiology and host interactions.
(S)-citramalyl-CoA lyase activity At A Glance
| GO ID | GO:0047777 |
|---|---|
| GO term | (S)-citramalyl-CoA lyase activity |
| Ontology | molecular_function |
| Synonym | (3S)-citramalyl-CoA lyase activity; citramalyl-CoA lyase activity; (+)-CMA-CoA lyase activity |
| Major function | Catalysis of (3S)-citramalyl-CoA = acetyl-CoA + pyruvate |
| Reaction type | Carbon-carbon bond cleavage (lyase) |
| Substrates | (3S)-citramalyl-CoA |
| Products | Acetyl-CoA and pyruvate |
| Pathway context | Bacterial itaconate degradation; autotrophic 3-hydroxypropionate cycle |
What Is GO:0047777?
According to the Gene Ontology, GO:0047777 (S)-citramalyl-CoA lyase activity is defined as the catalysis of the reaction: (3S)-citramalyl-CoA = acetyl-CoA + pyruvate. This is a lyase activity that breaks a carbon-carbon bond in (3S)-citramalyl-CoA to yield two central metabolites, acetyl-CoA and pyruvate. The term is also known by synonyms such as (3S)-citramalyl-CoA lyase activity, citramalyl-CoA lyase activity, and (+)-CMA-CoA lyase activity. It is a molecular_function term, meaning it describes what the enzyme does at the molecular level rather than a biological process or cellular component.
Why Is (S)-citramalyl-CoA lyase activity Important in Cell Biology?
GO:0047777 is important because it represents a metabolic node that links itaconate, a host-derived antimicrobial metabolite, to central carbon metabolism in bacteria. This activity enables pathogens such as Mycobacterium tuberculosis and Yersinia pestis to degrade itaconate and thereby resist its antimicrobial effects, contributing to pathogenicity. In environmental bacteria like Chloroflexus aurantiacus, the related enzyme supports autotrophic CO2 fixation through the 3-hydroxypropionate cycle. The activity is also relevant for metabolic engineering, as it can be harnessed or modified to improve itaconate production or degradation in biotechnological processes. Understanding its regulation and substrate specificity provides insights into microbial physiology, host-pathogen interactions, and potential drug targets.
• Enables bacterial pathogens to degrade itaconate, a host antimicrobial metabolite, supporting survival in macrophages.
• Contributes to the autotrophic 3-hydroxypropionate cycle for CO2 fixation in Chloroflexus aurantiacus.
• Represents a potential drug target because itaconate degradation is linked to virulence of Mycobacterium tuberculosis.
• Provides a metabolic route for converting itaconate-derived carbon into acetyl-CoA and pyruvate for energy and biosynthesis.
• Relevant for metabolic engineering of itaconate production and degradation pathways.
• Found in itaconate degradation gene clusters across Pseudomonas and other bacteria.
• Helps researchers understand carbon flux between host and pathogen during infection.
• Offers a model for studying lyase mechanism and substrate specificity.
• Can be studied using CRISPR-based gene editing to dissect gene function.
• Links microbial metabolism to host immune modulation via itaconate.
Molecular Mechanism of (S)-citramalyl-CoA lyase activity
Substrate recognition and binding
In simple terms: The enzyme grabs a specific molecule called (3S)-citramalyl-CoA and holds it in place.
The enzyme specifically binds (3S)-citramalyl-CoA, a CoA thioester, and positions it for cleavage. This specificity is crucial for its role in itaconate degradation, where itaconate is converted to (S)-citramalyl-CoA before cleavage. The enzyme is part of a gene cluster that includes other enzymes for itaconate metabolism, ensuring efficient substrate channeling.
Catalytic cleavage
In simple terms: The enzyme cuts the molecule into two smaller pieces: acetyl-CoA and pyruvate.
The lyase catalyzes the carbon-carbon bond cleavage of (3S)-citramalyl-CoA to produce acetyl-CoA and pyruvate. This reaction is a key step in the itaconate degradation pathway, allowing the carbon skeleton of itaconate to enter central metabolism. In Chloroflexus aurantiacus, a related enzyme (R-citramalyl-CoA lyase) performs an analogous reaction in the 3-hydroxypropionate cycle.
Cofactors and metal requirements
In simple terms: The enzyme may need helper molecules or metals to work, but details vary by organism.
The exact cofactor requirements for (S)-citramalyl-CoA lyase activity are not fully defined in the provided literature. However, related lyases in the itaconate degradation pathway often require divalent metal ions or specific cofactors for activity. Researchers studying this enzyme should experimentally determine its cofactor dependencies.
Regulation and pathway integration
In simple terms: The enzyme's activity is controlled by the cell's needs and is part of a larger metabolic pathway.
Expression of the (S)-citramalyl-CoA lyase gene is typically co-regulated with other genes in the itaconate degradation cluster, ensuring that the enzyme is produced when itaconate is available. In pathogenic bacteria, this pathway is induced during infection, allowing the pathogen to metabolize host-derived itaconate. The activity is integrated with central metabolism, as its products acetyl-CoA and pyruvate feed into the TCA cycle and other biosynthetic pathways.
Key Genes Involved in GO:0047777 (S)-citramalyl-CoA lyase activity
The following genes and proteins are associated with (S)-citramalyl-CoA lyase activity or its metabolic context, based on published literature.
| Gene | Major Role | Research Relevance |
|---|---|---|
| icl | Itaconate degradation cluster gene, encodes (S)-citramalyl-CoA lyase | Studied in Yersinia pestis for itaconate degradation and pathogenicity |
| iclR | Regulator of itaconate degradation genes | Controls expression of the itaconate degradation cluster |
| mct | Mesaconate utilization, related to itaconate metabolism | Studied in Burkholderia xenovorans for mesaconate utilization |
| fumC | Class I fumarase with mesaconase activity | Contributes to mesaconate utilization in Burkholderia xenovorans |
| prpD | Methylcitrate dehydratase, involved in propionate metabolism | Related to itaconate degradation in Pseudomonas |
| prpB | Methylisocitrate lyase, involved in propionate metabolism | Related to itaconate degradation in Pseudomonas |
| R-citramalyl-CoA lyase | Catalyzes similar reaction in 3-hydroxypropionate cycle | Studied in Chloroflexus aurantiacus for CO2 fixation |
| itaconate degradation cluster genes | Encode enzymes for itaconate degradation | Studied in Mycobacterium tuberculosis for itaconate dissimilation |
| thioesterase | Engineered for itaconate production via degradation scheme | Used in metabolic engineering for itaconate production |
| Pseudomonas itaconate degradation genes | Encode enzymes for methylsuccinate utilization | Studied in Pseudomonas for itaconate degradation |
| Burkholderia xenovorans mesaconase | Mesaconate utilization | Studied for mesaconate metabolism |
| Mycobacterium tuberculosis itaconate genes | Itaconate dissimilation | Studied for mechanism of action and dissimilation |
| Yersinia pestis icl | Itaconate degradation and pathogenicity | Studied for bacterial itaconate degradation |
| Chloroflexus aurantiacus R-citramalyl-CoA lyase | Autotrophic 3-hydroxypropionate cycle | Studied for CO2 fixation |
| Pseudomonas methylsuccinate genes | Methylsuccinate utilization | Studied for itaconate degradation gene cluster |
How Is (S)-citramalyl-CoA lyase activity Regulated?
The expression of genes encoding (S)-citramalyl-CoA lyase activity is regulated as part of the itaconate degradation gene cluster. In Yersinia pestis, the cluster is controlled by the regulator IclR, which responds to itaconate or its metabolites. In Mycobacterium tuberculosis, itaconate degradation genes are induced during infection, likely in response to host-derived itaconate. The activity itself may be regulated by substrate availability and feedback inhibition by products such as acetyl-CoA and pyruvate, though specific mechanisms require further study.
(S)-citramalyl-CoA lyase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| icl (Yersinia pestis) | Plague pathogenesis | Knockout in Y. pestis, infection model |
| Mycobacterium tuberculosis itaconate genes | Tuberculosis | Knockout in M. tuberculosis, macrophage infection |
| Pseudomonas itaconate genes | Opportunistic infections | Knockout in Pseudomonas, itaconate utilization assay |
| Burkholderia xenovorans mct/fumC | Mesaconate metabolism | Knockout in B. xenovorans, growth assay |
| Engineered thioesterase | Itaconate production | Overexpression in E. coli or yeast |
Tuberculosis and intracellular survival
Mycobacterium tuberculosis uses itaconate degradation, including (S)-citramalyl-CoA lyase activity, to metabolize host-derived itaconate and survive within macrophages. This activity is linked to the pathogen's ability to resist the antimicrobial effects of itaconate, making it a potential drug target.
Plague and bacterial pathogenicity
Yersinia pestis, the causative agent of plague, requires itaconate degradation for full pathogenicity. The (S)-citramalyl-CoA lyase activity is part of the itaconate degradation pathway that promotes bacterial survival in the host.
Metabolic engineering and biotechnology
The itaconate degradation pathway, including (S)-citramalyl-CoA lyase activity, is being explored for metabolic engineering of itaconate production. Engineering thioesterases to drive novel itaconate production via its degradation scheme highlights biotechnological relevance.
From (S)-citramalyl-CoA lyase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does knockout of icl reduce itaconate degradation? | CRISPR knockout in Yersinia pestis |
| Does point mutation in the active site abolish lyase activity? | CRISPR point mutation in the icl gene |
| Can knock-in of a tagged icl allow localization studies? | CRISPR knock-in of FLAG-tag in Y. pestis |
| Does overexpression of icl enhance itaconate utilization? | CRISPR overexpression in Pseudomonas |
| Does knockout of M. tuberculosis itaconate genes affect survival in macrophages? | CRISPR knockout in M. tuberculosis, macrophage infection |
| Can engineered thioesterase improve itaconate production? | Overexpression in E. coli |
How to Study the (S)-citramalyl-CoA lyase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Enzymatic assay | Lyase activity | Confirm gene function |
| CRISPR knockout | Gene essentiality | Test role in itaconate degradation |
| RNA-seq | Gene expression | Identify regulated genes |
| Proteomics | Protein abundance | Confirm enzyme production |
| Metabolomics | Metabolite levels | Measure pathway flux |
| Growth assays | Bacterial growth on itaconate | Phenotype of mutants |
| Macrophage infection | Intracellular survival | Virulence studies |
Enzymatic assays
Direct measurement of (S)-citramalyl-CoA lyase activity can be performed using spectrophotometric or chromatographic assays that detect the formation of acetyl-CoA or pyruvate from (3S)-citramalyl-CoA. Such assays are essential to confirm the function of candidate genes.
Gene knockout and complementation
CRISPR-based knockout of candidate genes followed by complementation with wild-type or mutant alleles can establish whether a gene encodes (S)-citramalyl-CoA lyase activity and whether the activity is required for itaconate degradation.
Transcriptomics and proteomics
RNA-seq and proteomics can reveal the expression of itaconate degradation genes under different conditions, such as during infection or in the presence of itaconate. This helps identify regulatory mechanisms and pathway integration.
Metabolomics
Metabolomic profiling can detect intermediates of the itaconate degradation pathway, including (3S)-citramalyl-CoA, acetyl-CoA, and pyruvate, providing evidence for flux through the (S)-citramalyl-CoA lyase step.
How CRISPR Can Be Used to Study GO:0047777 (S)-citramalyl-CoA lyase activity
Knockout
CRISPR knockout of genes encoding (S)-citramalyl-CoA lyase activity, such as icl in Yersinia pestis or homologous genes in Mycobacterium tuberculosis, can abolish itaconate degradation and reduce bacterial survival in macrophages. This approach directly tests the role of the activity in pathogenicity.
Point Mutation
CRISPR point mutation can be used to alter catalytic residues in the (S)-citramalyl-CoA lyase enzyme, allowing researchers to dissect the mechanism of substrate binding and cleavage. Such mutants can be tested for loss of enzymatic activity and reduced virulence.
Knock-in
CRISPR knock-in of epitope tags or fluorescent proteins into the endogenous locus enables visualization and purification of the (S)-citramalyl-CoA lyase enzyme, facilitating studies of its localization, interactions, and regulation.
Overexpression
CRISPR overexpression of the (S)-citramalyl-CoA lyase gene can enhance itaconate degradation or production, depending on the host. This is useful for metabolic engineering and for studying the effects of increased enzyme dosage.
How EDITGENE Supports (S)-citramalyl-CoA lyase activity Research
Researchers studying (S)-citramalyl-CoA lyase activity-related genes often need to determine whether a candidate gene is causally involved in itaconate degradation, pathogenicity, or metabolic engineering. EDITGENE provides CRISPR-based services to create precise knockout, point-mutation, knock-in, and overexpression cell models, as well as CRISPR library screening and bioinformatics support, to accelerate functional validation of genes in this pathway.
Contact EDITGENE today to design your custom CRISPR model for (S)-citramalyl-CoA lyase activity research.
Frequently Asked Questions About (S)-citramalyl-CoA lyase activity
What is (S)-citramalyl-CoA lyase activity?
It is an enzymatic activity defined by GO:0047777 that catalyzes the reaction (3S)-citramalyl-CoA = acetyl-CoA + pyruvate, a key step in bacterial itaconate degradation.
What genes are involved in (S)-citramalyl-CoA lyase activity?
Genes in the itaconate degradation cluster, such as icl in Yersinia pestis and homologous genes in Mycobacterium tuberculosis and Pseudomonas species, encode this activity.
What is the GO ID for (S)-citramalyl-CoA lyase activity?
The GO ID is GO:0047777.
Which organisms have (S)-citramalyl-CoA lyase activity?
It is found in bacteria such as Mycobacterium tuberculosis, Yersinia pestis, Pseudomonas species, Burkholderia xenovorans, and Chloroflexus aurantiacus.
How is (S)-citramalyl-CoA lyase activity related to disease?
It supports bacterial survival in hosts by degrading itaconate, contributing to pathogenicity of Mycobacterium tuberculosis and Yersinia pestis.
What is the reaction catalyzed by (S)-citramalyl-CoA lyase?
The enzyme cleaves (3S)-citramalyl-CoA into acetyl-CoA and pyruvate [QuickGO].
Can CRISPR be used to study (S)-citramalyl-CoA lyase activity?
Yes, CRISPR knockout, point mutation, knock-in, and overexpression can be used to dissect gene function and regulation.
What are the products of (S)-citramalyl-CoA lyase activity?
The products are acetyl-CoA and pyruvate, both central metabolites [QuickGO].
Is (S)-citramalyl-CoA lyase activity involved in CO2 fixation?
A related enzyme, R-citramalyl-CoA lyase, participates in the autotrophic 3-hydroxypropionate cycle in Chloroflexus aurantiacus.
How can I study (S)-citramalyl-CoA lyase activity in the lab?
Enzymatic assays, CRISPR gene editing, metabolomics, and transcriptomics are common approaches.
Conclusion
GO:0047777 (S)-citramalyl-CoA lyase activity is a molecular function that plays a critical role in bacterial itaconate degradation and related metabolic pathways. Its importance spans pathogenicity, host-pathogen interactions, and biotechnological applications. By leveraging CRISPR-based gene editing and multi-omics approaches, researchers can uncover the precise mechanisms and regulatory networks controlling this activity, potentially leading to new therapeutic or engineering strategies.
References
- 1. Priya M et al.. 2025. Itaconate mechanism of action and dissimilation in Mycobacterium tuberculosis.. Proc Natl Acad Sci U S A 122(4):e2423114122 PMID: 39841148
- 2. Kronen M et al.. 2015. Mesaconase Activity of Class I Fumarase Contributes to Mesaconate Utilization by Burkholderia xenovorans.. Appl Environ Microbiol 81(16):5632-8 PMID: 26070669
- 3. Sasikaran J et al.. 2014. Bacterial itaconate degradation promotes pathogenicity.. Nat Chem Biol 10(5):371-7 PMID: 24657929
- 4. Wang RS et al.. 2024. Engineering thioesterase as a driving force for novel itaconate production via its degradation scheme.. Metab Eng Commun 19:e00246 PMID: 39224858
- 5. Friedmann S et al.. 2007. Properties of R-citramalyl-coenzyme A lyase and its role in the autotrophic 3-hydroxypropionate cycle of Chloroflexus aurantiacus.. J Bacteriol 189(7):2906-14 PMID: 17259315
- 6. Gonner L et al.. 2025. Pseudomonadal itaconate degradation gene cluster encodes enzymes for methylsuccinate utilization.. Commun Biol 8(1):1099 PMID: 40707647
