GO:0019171 (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0019171 describes the dehydration of a (3R)-hydroxyacyl-[acyl-carrier-protein] to a (2E)-enoyl-[acyl-carrier-protein] plus water, a core step in fatty acid biosynthesis.
• The reaction is catalyzed by FabZ-type dehydratases in type II fatty acid synthase (FAS-II) systems, including the Pseudomonas aeruginosa FabZ enzyme whose structure was solved by Kimber et al..
• In Mycobacterium tuberculosis, the FAS-II dehydratase activity is associated with HadBD, a singular structure with a unique function.
• The same dehydratase chemistry is shared by peroxisomal enzymes such as D-bifunctional protein, where mutations cause D-bifunctional protein deficiency [5,6].
• Plant and mammalian peroxisomal enoyl-CoA hydratase 2 enzymes catalyze related hydratase/dehydratase reactions in fatty acid degradation and bile acid synthesis [7,8].
• GO:0019171 is a molecular_function term and is a target for antibacterial and antituberculosis drug discovery [2,3].
Description
GO:0019171, (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity, is a molecular_function term describing the catalytic removal of water from a (3R)-hydroxyacyl-[acyl-carrier-protein] to form a (2E)-enoyl-[acyl-carrier-protein]. This dehydration step is essential in type II fatty acid synthase (FAS-II) systems, where each cycle of chain elongation requires the formation of a trans-2-enoyl intermediate before reduction. The enzyme FabZ from Pseudomonas aeruginosa is a structurally characterized example of this activity, and its crystal structure revealed the fold and active-site architecture of the dehydratase. In Mycobacterium tuberculosis, the FAS-II system relies on a distinct dehydratase, HadBD, which has a singular structure and a unique function in the pathway. The same chemical transformation is also catalyzed by peroxisomal enzymes such as D-bifunctional protein, where the dehydratase domain participates in fatty acid beta-oxidation [5,6]. Researchers study GO:0019171 because it is both a central node in lipid metabolism and a validated target for antibacterial and antituberculosis compounds [2,3].
(3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity At A Glance
| GO ID | GO:0019171 |
|---|---|
| GO term | (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity |
| Ontology | molecular_function |
| Synonym | 3-hydroxyacyl-ACP dehydratase activity; 3-hydroxyacyl-[acyl-carrier protein] dehydratase activity; 3-hydroxyacyl-[acyl-carrier-protein] dehydratase activity; (3R)-3-hydroxyacyl-[acyl-carrier-protein] dehydratase activity |
| Major function | Catalysis of the dehydration of a (3R)-hydroxyacyl-[acyl-carrier-protein] to a (2E)-enoyl-[acyl-carrier-protein] plus water |
| Representative enzyme | FabZ from Pseudomonas aeruginosa, a structurally characterized dehydratase |
| Related enzyme | HadBD from Mycobacterium tuberculosis, a FAS-II dehydratase with a unique structure |
| Pathway context | Type II fatty acid synthase (FAS-II) elongation cycle [1,4] |
| Disease relevance | D-bifunctional protein deficiency involves a peroxisomal dehydratase domain [5,6] |
What Is GO:0019171?
GO:0019171 is defined by QuickGO as the catalysis of the reaction: a (3R)-hydroxyacyl-[acyl-carrier-protein] = a (2E)-enoyl-[acyl-carrier-protein] + H2O. In other words, the enzyme removes a water molecule from a 3R-hydroxyacyl chain that is attached to an acyl-carrier protein, generating a trans-2-enoyl product. This is a dehydration reaction and is a standard step in fatty acid biosynthesis and related lipid pathways.
Why Is (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity Important in Cell Biology?
GO:0019171 is important because it represents a chemically essential step in fatty acid biosynthesis and related lipid metabolic pathways. In bacteria, the dehydratase reaction is required for the FAS-II elongation cycle, and the enzyme FabZ is a validated target for antibacterial compounds [1,2,3]. In Mycobacterium tuberculosis, the HadBD dehydratase is a structurally distinct enzyme that performs this activity, making it an attractive target for antituberculosis drug discovery. In humans, peroxisomal enzymes with dehydratase activity, such as D-bifunctional protein, are required for fatty acid beta-oxidation, and mutations in the corresponding gene cause D-bifunctional protein deficiency [5,6]. Thus, understanding GO:0019171 informs both antimicrobial development and inherited metabolic disease research.
• It is a core catalytic step in the type II fatty acid synthase (FAS-II) elongation cycle.
• The Pseudomonas aeruginosa FabZ structure provides a template for understanding dehydratase mechanism and inhibitor design.
• In Mycobacterium tuberculosis, HadBD is a unique FAS-II dehydratase with a singular structure.
• Phenotypic screens for M. tuberculosis FAS-II inhibitors can lack specificity, making target-based knowledge of GO:0019171 important.
• New antituberculosis compounds have been developed against FAS-II-related targets, including dehydratase-associated chemistry.
• D-bifunctional protein deficiency is caused by mutations affecting a peroxisomal enzyme with dehydratase activity [5,6].
• Peroxisomal enoyl-CoA hydratase 2 enzymes participate in degradation of even cis-unsaturated fatty acids in plants.
• Rat peroxisomal multifunctional enzyme 2 has a hydratase reaction involved in bile acid synthesis.
• The term links molecular function to lipid metabolism, membrane biogenesis, and energy homeostasis [1,4].
• It provides a defined target for CRISPR knockout, point-mutation, and inhibitor studies in microbial and mammalian systems [1,4,5].
Molecular Mechanism of (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity
Substrate recognition and acyl-carrier protein binding
In simple terms: The enzyme grabs a fatty acid chain that is carried by a small protein called acyl-carrier protein.
The substrate for GO:0019171 is a (3R)-hydroxyacyl-[acyl-carrier-protein], in which the growing fatty acid chain is covalently attached to an acyl-carrier protein. The dehydratase must recognize this acyl-carrier protein-bound substrate and position the 3R-hydroxyacyl chain in the active site for catalysis. In the FAS-II system, this step follows the reduction of a 3-ketoacyl intermediate and precedes the final reduction of the enoyl product [1,4].
Catalytic dehydration to form the trans-2-enoyl product
In simple terms: The enzyme removes a water molecule from the fatty acid chain, creating a double bond.
The catalytic reaction of GO:0019171 is the dehydration of a (3R)-hydroxyacyl-[acyl-carrier-protein] to a (2E)-enoyl-[acyl-carrier-protein] plus water. This creates a trans double bond at the 2-position of the acyl chain, which is the substrate for the subsequent enoyl reductase step in FAS-II. The reaction is reversible in principle, but in the context of fatty acid biosynthesis it proceeds in the dehydrating direction to drive chain elongation.
Structural basis: the FabZ fold
In simple terms: The enzyme has a specific three-dimensional shape that makes the reaction possible.
The crystal structure of FabZ from Pseudomonas aeruginosa revealed the structural basis for (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity. This structure showed the active-site architecture and provided insight into how the enzyme binds the acyl-carrier protein substrate and catalyzes dehydration. The FabZ structure is a reference for understanding the mechanism of GO:0019171 and for structure-based inhibitor design.
HadBD: a distinct FAS-II dehydratase in Mycobacterium tuberculosis
In simple terms: Tuberculosis bacteria use a different-looking enzyme to do the same dehydration step.
In Mycobacterium tuberculosis, the FAS-II system uses HadBD, a dehydratase with a singular structure and a unique function. This enzyme performs the (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity within the mycobacterial FAS-II pathway. The distinct structure of HadBD makes it an attractive target for antituberculosis drug development.
Related peroxisomal dehydratase activities
In simple terms: Similar dehydration reactions happen in human and plant peroxisomes during fat breakdown.
D-bifunctional protein is a peroxisomal enzyme with dehydratase activity, and mutations in its gene cause D-bifunctional protein deficiency [5,6]. The molecular basis of D-bifunctional protein deficiency type III has been linked to specific mutations affecting the dehydratase function. In plants, a monofunctional peroxisomal enoyl-CoA hydratase 2 participates in the degradation of even cis-unsaturated fatty acids, and rat peroxisomal multifunctional enzyme 2 has a hydratase reaction involved in bile acid synthesis. These enzymes illustrate the broader biochemical context of dehydratase chemistry related to GO:0019171.
Key Genes Involved in GO:0019171 (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity
The following genes and proteins are experimentally linked to (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity or to related dehydratase reactions in fatty acid metabolism.
| Gene | Major Role | Research Relevance |
|---|---|---|
| fabZ (Pseudomonas aeruginosa) | Encodes the (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase FabZ | Structurally characterized enzyme for GO:0019171; model for mechanism and inhibitor design |
| hadB (Mycobacterium tuberculosis) | Component of the HadBD dehydratase in FAS-II | Unique FAS-II dehydratase structure and function; drug target |
| hadD (Mycobacterium tuberculosis) | Component of the HadBD dehydratase in FAS-II | Unique FAS-II dehydratase structure and function; drug target |
| hsd17b4 (human) | Encodes D-bifunctional protein with dehydratase activity | Mutations cause D-bifunctional protein deficiency [5,6] |
| HSD17B4 (human) | Peroxisomal multifunctional enzyme with dehydratase domain | Genotype-phenotype analysis of D-bifunctional protein deficiency |
| ech2 (Arabidopsis thaliana) | Monofunctional peroxisomal enoyl-CoA hydratase 2 | Degradation of even cis-unsaturated fatty acids |
| MFP2 (rat) | Peroxisomal multifunctional enzyme 2 with hydratase activity | Role of hydratase reaction in bile acid synthesis |
| fabA (Escherichia coli) | Dehydratase/isomerase in FAS-II | Model for bacterial dehydratase mechanism (contextual) |
| fabZ (Escherichia coli) | Dehydratase in FAS-II | Model for bacterial dehydratase mechanism (contextual) |
| inhA (Mycobacterium tuberculosis) | Enoyl reductase in FAS-II | Target of antituberculosis drugs; pathway context |
| kasA (Mycobacterium tuberculosis) | Ketoacyl synthase in FAS-II | Pathway context for dehydratase step |
| kasB (Mycobacterium tuberculosis) | Ketoacyl synthase in FAS-II | Pathway context for dehydratase step |
| acpM (Mycobacterium tuberculosis) | Acyl-carrier protein in FAS-II | Provides the acyl-carrier protein substrate for GO:0019171 |
| fabD (Mycobacterium tuberculosis) | Malonyl-CoA:ACP transacylase | Pathway context for FAS-II |
| fabH (Mycobacterium tuberculosis) | Ketoacyl synthase in FAS-II | Pathway context for FAS-II |
| fabI (Mycobacterium tuberculosis) | Enoyl reductase in FAS-II | Pathway context for FAS-II |
| fabZ (Mycobacterium tuberculosis) | Dehydratase in FAS-II | Related dehydratase in mycobacteria |
How Is (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity Regulated?
The regulation of GO:0019171 is primarily contextual to the FAS-II pathway and peroxisomal fatty acid oxidation. In Mycobacterium tuberculosis, phenotypic screens for FAS-II inhibitors can lack specificity, indicating that pathway-level regulation and target engagement are complex. The HadBD dehydratase is a distinct component of the mycobacterial FAS-II system, and its unique structure suggests specialized regulation within the pathway. In peroxisomes, D-bifunctional protein deficiency results from mutations that impair dehydratase function, and the clinical phenotype depends on the specific mutation and residual activity [5,6]. Plant peroxisomal enoyl-CoA hydratase 2 is involved in degradation of even cis-unsaturated fatty acids, linking its regulation to lipid mobilization. Rat peroxisomal multifunctional enzyme 2 hydratase activity participates in bile acid synthesis, connecting the reaction to cholesterol metabolism.
(3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| HSD17B4 | D-bifunctional protein deficiency [5,6] | Patient-derived fibroblasts or CRISPR knock-in of patient mutations |
| hadB/hadD | Mycobacterium tuberculosis FAS-II function and drug target | Mycobacterial knockout and conditional depletion strains |
| fabZ (Pseudomonas aeruginosa) | Bacterial fatty acid biosynthesis and antibacterial target | Recombinant enzyme assays and bacterial knockout |
| fabZ (Escherichia coli) | Model bacterial FAS-II dehydratase | Genetic knockout and complementation |
| ech2 (Arabidopsis thaliana) | Peroxisomal fatty acid degradation | Plant knockout lines and lipid profiling |
D-bifunctional protein deficiency
D-bifunctional protein deficiency is a peroxisomal disorder caused by mutations in the gene encoding D-bifunctional protein, which has dehydratase activity [5,6]. The molecular basis of type III deficiency has been studied in detail, and structure-based genotype-phenotype analysis has linked specific mutations to loss of dehydratase function [5,6]. This disease demonstrates that impaired (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase-related chemistry in peroxisomes has severe clinical consequences [5,6].
Tuberculosis and FAS-II targeting
Mycobacterium tuberculosis relies on the FAS-II system for mycolic acid biosynthesis, and the HadBD dehydratase performs a unique function in this pathway. Phenotypic screens for FAS-II inhibitors can lack specificity, highlighting the need for target-based approaches against enzymes such as HadBD. New antituberculosis compounds have been developed against FAS-II-related targets, supporting the therapeutic relevance of dehydratase chemistry.
Bacterial infections and antibacterial discovery
The Pseudomonas aeruginosa FabZ enzyme is a structurally characterized (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase, and its active site is a potential target for antibacterial agents. Because the dehydratase step is essential in bacterial FAS-II, inhibitors of GO:0019171 could block fatty acid biosynthesis and bacterial growth [1,3].
From (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Is FabZ essential for bacterial growth? | CRISPR or homologous recombination knockout of fabZ in Pseudomonas aeruginosa |
| Does a patient mutation impair dehydratase activity? | CRISPR knock-in of the mutation into HSD17B4 in human cells [5,6] |
| Can a compound inhibit the dehydratase step? | Recombinant FabZ enzyme assay with candidate inhibitors |
| What is the role of HadBD in M. tuberculosis FAS-II? | Conditional knockdown or knockout of hadB/hadD in mycobacteria |
| Does overexpression of a dehydratase alter lipid profiles? | CRISPR overexpression of fabZ or HSD17B4 in cell lines [1,5] |
| How does loss of peroxisomal dehydratase affect bile acid synthesis? | Knockout of MFP2 in rat or mouse models |
How to Study the (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Recombinant enzyme assay | Dehydratase catalytic activity | Testing FabZ inhibitors and mutants |
| X-ray crystallography | Three-dimensional protein structure | Active-site analysis of FabZ and HadBD [1,4] |
| Phenotypic screening | Growth inhibition of bacteria | FAS-II inhibitor discovery |
| Target-based screening | Inhibition of purified enzyme | Specific dehydratase inhibitor identification |
| Lipidomics | Fatty acid composition changes | Evaluating pathway flux after genetic perturbation [4,7] |
| Bile acid profiling | Bile acid synthesis | Assessing peroxisomal hydratase function |
| CRISPR knockout | Loss-of-function phenotype | Testing gene essentiality in bacteria and cells [1,4] |
| CRISPR knock-in | Mutant protein function | Modeling patient mutations in HSD17B4 [5,6] |
Enzyme activity assays
Direct measurement of (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity can be performed using recombinant enzymes such as FabZ and synthetic acyl-carrier protein substrates. These assays monitor the formation of the (2E)-enoyl-[acyl-carrier-protein] product, often by spectrophotometric or chromatographic methods. Such assays are used to test inhibitors and to characterize mutant enzymes.
Structural biology
X-ray crystallography of FabZ from Pseudomonas aeruginosa revealed the fold and active-site architecture of the dehydratase. Structural studies of HadBD from Mycobacterium tuberculosis showed a singular structure for this unique FAS-II enzyme. These structures guide mechanistic understanding and structure-based inhibitor design [1,4].
Genetic and phenotypic screens
Phenotypic screens for FAS-II inhibitors can be used to identify compounds that target the pathway, but they may lack specificity. Target-based screens against purified dehydratase enzymes complement phenotypic approaches [1,3]. Antituberculosis compounds have been evaluated in such screens, linking chemical matter to FAS-II function.
Lipidomics and metabolic profiling
Lipid profiling can reveal changes in fatty acid composition when dehydratase activity is altered [4,7]. In plants, analysis of even cis-unsaturated fatty acid degradation requires monitoring of peroxisomal enzyme activity. In mammals, bile acid synthesis can be assessed to evaluate the role of peroxisomal hydratase reactions.
How CRISPR Can Be Used to Study GO:0019171 (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity
Knockout
CRISPR knockout of fabZ or hadB/hadD can be used to test whether the dehydratase is essential for bacterial growth and FAS-II function [1,4]. In human cells, knockout of HSD17B4 can model loss of peroxisomal dehydratase activity and reveal metabolic consequences [5,6]. Knockout studies provide causal evidence linking GO:0019171 to cellular phenotypes.
Point Mutation
CRISPR point mutation can introduce specific active-site residues or patient-derived mutations into dehydratase genes to test their effect on catalysis [1,5]. For example, mutations identified in D-bifunctional protein deficiency can be modeled to assess residual dehydratase activity [5,6]. Point-mutation models help distinguish loss-of-function from structural effects.
Knock-in
CRISPR knock-in can insert epitope tags or fluorescent reporters into endogenous dehydratase genes to study localization and interactions [1,4]. Knock-in of disease-associated mutations into HSD17B4 allows study of genotype-phenotype relationships [5,6]. Tagged knock-in lines enable proteomic and imaging analyses of the dehydratase.
Overexpression
CRISPR overexpression of fabZ or HSD17B4 can increase dehydratase activity and reveal effects on lipid metabolism [1,5]. Overexpression models are useful for producing recombinant enzyme for structural and biochemical studies. They can also test whether increased dehydratase activity alters pathway flux or drug sensitivity [1,3].
How EDITGENE Supports (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity Research
Researchers studying (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity-related genes often need to determine whether a candidate gene is causally involved in fatty acid metabolism, drug response, or disease. EDITGENE provides CRISPR-based cell models and screening services to enable these investigations.
Contact EDITGENE today to design your custom CRISPR model for (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity research.
Frequently Asked Questions About (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity
What is GO:0019171?
GO:0019171 is the Gene Ontology molecular_function term for (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity, which catalyzes the dehydration of a (3R)-hydroxyacyl-[acyl-carrier-protein] to a (2E)-enoyl-[acyl-carrier-protein] plus water.
What reaction does (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase catalyze?
It removes water from a (3R)-hydroxyacyl-[acyl-carrier-protein] to form a (2E)-enoyl-[acyl-carrier-protein], a step in fatty acid biosynthesis.
What genes are involved in (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity?
Genes include fabZ in Pseudomonas aeruginosa and Escherichia coli, hadB and hadD in Mycobacterium tuberculosis, and HSD17B4 in humans [1,4,5,6].
Which enzyme structure represents GO:0019171?
The crystal structure of FabZ from Pseudomonas aeruginosa is a representative structure for this activity.
How is HadBD related to GO:0019171?
HadBD is a Mycobacterium tuberculosis FAS-II dehydratase with a unique structure that performs the (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase reaction.
What diseases are linked to dehydratase activity?
D-bifunctional protein deficiency is caused by mutations in a peroxisomal enzyme with dehydratase activity, and FAS-II dehydratases are targets for tuberculosis and bacterial infections [4,5,6].
How can I study GO:0019171 in the lab?
Recombinant enzyme assays, X-ray crystallography, CRISPR knockout or knock-in, and lipidomics are common approaches [1,4,5].
Is (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase a drug target?
Yes, bacterial and mycobacterial dehydratases are targets for antibacterial and antituberculosis drug discovery [1,2,3,4].
What is the difference between FabZ and HadBD?
FabZ is a well-characterized bacterial dehydratase, while HadBD is a structurally distinct FAS-II dehydratase in Mycobacterium tuberculosis [1,4].
Can CRISPR be used to model dehydratase deficiency?
Yes, CRISPR knockout or knock-in of HSD17B4 can model D-bifunctional protein deficiency and test mutation effects [5,6].
Conclusion
GO:0019171, (3R)-hydroxyacyl-[acyl-carrier-protein] dehydratase activity, is a central molecular function in fatty acid biosynthesis and related lipid pathways. Its structural and mechanistic characterization, exemplified by FabZ and HadBD, provides a foundation for antibacterial and antituberculosis drug discovery [1,4]. In humans, peroxisomal enzymes with dehydratase activity are linked to D-bifunctional protein deficiency, underscoring the clinical importance of this chemistry [5,6]. CRISPR-based models and biochemical assays enable researchers to dissect the function of GO:0019171 in health and disease.
References
- 1. Kimber MS et al.. 2004. The structure of (3R)-hydroxyacyl-acyl carrier protein dehydratase (FabZ) from Pseudomonas aeruginosa.. J Biol Chem 279(50):52593-602 PMID: 15371447
- 2. Pretelín-Castillo G et al.. 2021. (2Z)-3-Hydroxy-3-(4-R-Phenyl)-Prop-2-Enedithioic Acids as New Antituberculosis Compounds.. Infect Drug Resist 14:4323-4332 PMID: 34707377
- 3. Grzegorzewicz AE et al.. 2020. Lack of Specificity of Phenotypic Screens for Inhibitors of the Mycobacterium tuberculosis FAS-II System.. Antimicrob Agents Chemother 65(1) PMID: 33139282
- 4. Bories P et al.. 2024. HadBD dehydratase from Mycobacterium tuberculosis fatty acid synthase type II: A singular structure for a unique function.. Protein Sci 33(4):e4964 PMID: 38501584
- 5. Mehtälä ML et al.. 2013. On the molecular basis of D-bifunctional protein deficiency type III.. PLoS One 8(1):e53688 PMID: 23308274
- 6. Ferdinandusse S et al.. 2006. Mutational spectrum of D-bifunctional protein deficiency and structure-based genotype-phenotype analysis.. Am J Hum Genet 78(1):112-24 PMID: 16385454
- 7. Goepfert S et al.. 2006. Identification and functional characterization of a monofunctional peroxisomal enoyl-CoA hydratase 2 that participates in the degradation of even cis-unsaturated fatty acids in Arabidopsis thaliana.. J Biol Chem 281(47):35894-903 PMID: 16982622
- 8. Qin YM et al.. 1997. Recombinant 2-enoyl-CoA hydratase derived from rat peroxisomal multifunctional enzyme 2: role of the hydratase reaction in bile acid synthesis.. Biochem J 328 ( Pt 2)(Pt 2):377-82 PMID: 9371691
