GO:0017057 6-phosphogluconolactonase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0017057 defines the enzymatic activity that hydrolyzes 6-O-phosphono-D-glucono-1,5-lactone to 6-phospho-D-gluconate, a critical step in the pentose phosphate pathway.
• 6-phosphogluconolactonase (6PGL) prevents the accumulation of the reactive lactone intermediate, which can otherwise modify proteins and impair cellular function [2,3].
• In some organisms, 6PGL is fused to glucose-6-phosphate dehydrogenase (G6PD), forming a bifunctional enzyme, as seen in Plasmodium species and murine hexose-6-phosphate dehydrogenase [5,7,8].
• Loss of 6PGL activity leads to metabolic disruption and is linked to disease; for example, in Leishmania donovani, the enzyme is crucial for growth and host infection.
• Hyperpolarized δ-[1-13C]gluconolactone has been used to image 6PGL activity in brain tumors in vivo, highlighting its potential as a metabolic imaging biomarker.
• Plant 6PGL isoforms in peroxisomes show potential redundancy, indicating complex regulation of the oxidative pentose phosphate pathway.
Description
6-phosphogluconolactonase (6PGL) activity, encoded by GO:0017057, catalyzes the hydrolysis of 6-O-phosphono-D-glucono-1,5-lactone to 6-phospho-D-gluconate and H+. This reaction is the second step of the oxidative phase of the pentose phosphate pathway (PPP), following the glucose-6-phosphate dehydrogenase (G6PD)-catalyzed production of the lactone. By rapidly removing the lactone intermediate, 6PGL prevents its spontaneous hydrolysis and the accumulation of reactive species that can damage cellular components [2,3]. The enzyme is therefore essential for maintaining flux through the PPP, which supplies NADPH and ribose-5-phosphate for biosynthesis and antioxidant defense. Research on 6PGL spans diverse organisms, from bacteria to humans, and has revealed both canonical and fused forms of the enzyme. In Escherichia coli, 6PGL is a standalone protein whose activity is inhibited by oxidative modification and aggregation. In contrast, Plasmodium falciparum and Plasmodium vivax express a bifunctional enzyme where 6PGL is fused to G6PD, making it an attractive drug target [7,8]. Murine hexose-6-phosphate dehydrogenase also exhibits intrinsic 6PGL activity, suggesting evolutionary adaptations. In plants, peroxisomal 6PGL isoforms contribute to NADPH production for redox metabolism. Given its central role in metabolism, 6PGL is increasingly recognized as a potential therapeutic target and biomarker. For instance, hyperpolarized δ-[1-13C]gluconolactone imaging has been used to non-invasively assess 6PGL activity in brain tumors. In Leishmania donovani, 6PGL is critical for growth and host infection, underscoring its importance in pathogen biology. This article provides a comprehensive overview of GO:0017057, covering its definition, mechanism, key genes, disease relevance, and research methodologies, with a focus on CRISPR-based models for functional studies.
6-phosphogluconolactonase activity At A Glance
| GO ID | GO:0017057 |
|---|---|
| GO term | 6-phosphogluconolactonase activity |
| Ontology | molecular_function |
| Synonym | 6-PGL, 6-phospho-D-glucono-1,5-lactone lactonohydrolase activity, phosphogluconolactonase activity |
| Major function | Catalyzes the hydrolysis of 6-O-phosphono-D-glucono-1,5-lactone to 6-phospho-D-gluconate and H+ |
| Reaction | 6-O-phosphono-D-glucono-1,5-lactone + H2O = 6-phospho-D-gluconate + H+ |
| Pathway | Pentose phosphate pathway (oxidative phase) |
| EC number | 3.1.1.31 |
| Substrate | 6-O-phosphono-D-glucono-1,5-lactone |
| Product | 6-phospho-D-gluconate |
What Is GO:0017057?
GO:0017057, 6-phosphogluconolactonase activity, is a molecular function defined by the catalysis of the reaction: 6-O-phosphono-D-glucono-1,5-lactone + H2O = 6-phospho-D-gluconate + H+. This activity is synonymous with 6-PGL, 6-phospho-D-glucono-1,5-lactone lactonohydrolase activity, and phosphogluconolactonase activity. It is a hydrolase that acts on the lactone intermediate of the pentose phosphate pathway, ensuring efficient carbon flux and preventing the buildup of a reactive metabolite.
Why Is 6-phosphogluconolactonase activity Important in Cell Biology?
6-phosphogluconolactonase activity is essential for the proper functioning of the pentose phosphate pathway, which provides NADPH and ribose-5-phosphate for reductive biosynthesis and nucleotide synthesis. By efficiently hydrolyzing the lactone intermediate, 6PGL prevents the accumulation of a reactive electrophile that can modify proteins and cause cellular damage [2,3]. Dysregulation of this activity has been implicated in various diseases, including cancer, where metabolic reprogramming often upregulates the PPP, and in parasitic infections, where the enzyme is vital for pathogen survival [4,7,8]. Understanding 6PGL is therefore crucial for developing targeted therapies and diagnostic tools.
• Maintains flux through the pentose phosphate pathway, supporting NADPH production and antioxidant defense.
• Prevents accumulation of the reactive lactone intermediate, which can cause protein modification and aggregation.
• Serves as a potential drug target in parasites such as Plasmodium falciparum and Leishmania donovani [4,7,8].
• Enables non-invasive imaging of tumor metabolism using hyperpolarized δ-[1-13C]gluconolactone.
• Plays a role in plant peroxisomal redox metabolism, with isoforms showing potential redundancy.
• Fused forms with G6PD in some organisms provide a model for bifunctional enzyme evolution [5,7,8].
• Its activity can be modulated by oxidative stress, linking it to cellular stress responses.
• Deficiency or inhibition leads to metabolic disruption and growth defects in model organisms [3,4].
Molecular Mechanism of 6-phosphogluconolactonase activity
Substrate Binding and Catalysis
In simple terms: The enzyme grabs the lactone molecule and uses water to break it open, turning it into a useful product.
6-phosphogluconolactonase binds its substrate, 6-O-phosphono-D-glucono-1,5-lactone, and catalyzes its hydrolysis to 6-phospho-D-gluconate and H+. This reaction is essential because the lactone can spontaneously hydrolyze, but the enzyme accelerates the process and prevents the lactone from reacting with cellular nucleophiles. The catalytic mechanism likely involves general acid-base chemistry, although specific residues have not been fully characterized in all organisms. In E. coli, oxidative modification of specific amino acids by peroxyl radicals inhibits enzyme activity and promotes aggregation, indicating that catalytic residues are vulnerable to oxidative damage.
Fused and Standalone Enzyme Architectures
In simple terms: In some organisms, the enzyme is part of a larger protein that does two jobs, while in others it works alone.
In many organisms, 6PGL exists as a standalone enzyme, such as in E. coli and Arabidopsis [2,6]. However, in Plasmodium falciparum and Plasmodium vivax, the enzyme is fused to glucose-6-phosphate dehydrogenase (G6PD), forming a bifunctional protein where the 6PGL domain follows the G6PD domain [7,8]. This fusion may facilitate substrate channeling, as the product of G6PD (the lactone) is directly handed to the 6PGL active site. Murine hexose-6-phosphate dehydrogenase also exhibits 6PGL activity, suggesting that bifunctional architectures are not limited to parasites.
Role in the Pentose Phosphate Pathway
In simple terms: This enzyme is the second step in a pathway that makes building blocks and reducing power for cells.
6PGL catalyzes the second step of the oxidative phase of the pentose phosphate pathway, immediately following the G6PD-catalyzed oxidation of glucose-6-phosphate to 6-phosphogluconolactone. The product, 6-phospho-D-gluconate, is then further metabolized by 6-phosphogluconate dehydrogenase to produce ribulose-5-phosphate and NADPH. Thus, 6PGL is critical for maintaining the flux of carbon through the PPP, which is essential for nucleotide biosynthesis and antioxidant defense. In Leishmania donovani, loss of 6PGL impairs growth and host infection, highlighting its importance in pathogen metabolism.
Regulation and Post-translational Modifications
In simple terms: The enzyme's activity can be turned up or down by chemical changes or stress conditions.
6PGL activity can be regulated by oxidative stress. In E. coli, peroxyl radicals modify specific amino acids, leading to enzyme aggregation and inhibition. This suggests that under oxidative stress, 6PGL may be a target for damage, potentially affecting PPP flux. In Arabidopsis, peroxisomal 6PGL isoforms may be regulated at the level of gene expression or protein targeting, as analysis of potential redundancy indicates that multiple isoforms contribute to activity. However, specific transcriptional regulators or signaling pathways controlling 6PGL expression remain largely uncharacterized.
Inhibitors and Chemical Probes
In simple terms: Scientists have found molecules that can block this enzyme, which could be developed into drugs.
Inhibitor studies on the Plasmodium vivax G6PD-6PGL bifunctional enzyme have identified compounds that target the 6PGL domain, suggesting that this activity is druggable. Similarly, the essential role of 6PGL in Leishmania donovani growth and infection makes it a potential target for antiparasitic chemotherapy. In cancer, hyperpolarized δ-[1-13C]gluconolactone has been used to image 6PGL activity, providing a non-invasive method to assess enzyme levels and potentially evaluate inhibitor efficacy.
Key Genes Involved in GO:0017057 6-phosphogluconolactonase activity
The following genes and proteins are directly associated with 6-phosphogluconolactonase activity or its fused forms across various organisms.
| Gene | Major Role | Research Relevance |
|---|---|---|
| PGLS (human) | Encodes 6-phosphogluconolactonase | Studied for its role in PPP and potential as a cancer target |
| pgl (E. coli) | Standalone 6PGL | Model for oxidative stress-induced inhibition and aggregation |
| G6PD-6PGL (P. falciparum) | Bifunctional enzyme with G6PD and 6PGL activities | Potential drug target for malaria |
| G6PD-6PGL (P. vivax) | Bifunctional enzyme | Characterized for inhibitor studies |
| H6PD (mouse) | Hexose-6-phosphate dehydrogenase with 6PGL activity | Bifunctional enzyme model |
| AtPGL1-3 (Arabidopsis) | Peroxisomal 6PGL isoforms | Studied for redundancy in peroxisomal NADPH production |
| LdPGL (Leishmania donovani) | 6PGL essential for growth and infection | Drug target for leishmaniasis |
| PGLS (other mammals) | 6PGL homologs | Comparative studies of PPP regulation |
| DevB (Vibrio cholerae) | 6PGL homolog | Potential role in pathogenesis |
| PGL (yeast) | 6PGL in Saccharomyces cerevisiae | Model for PPP genetics |
| PGL (Drosophila) | 6PGL in fruit fly | Developmental studies |
| PGL (zebrafish) | 6PGL in Danio rerio | Vertebrate model for PPP |
| PGL (C. elegans) | 6PGL in nematode | Aging and stress studies |
| G6PD (human) | Glucose-6-phosphate dehydrogenase | Produces lactone substrate for 6PGL |
| 6PGD (human) | 6-phosphogluconate dehydrogenase | Downstream enzyme in PPP |
| PGLS (mouse) | Murine 6PGL | Knockout models for metabolic studies |
| PGLS (rat) | Rat 6PGL | Biochemical characterization |
| PGLS (bovine) | Bovine 6PGL | Structural studies |
How Is 6-phosphogluconolactonase activity Regulated?
6-phosphogluconolactonase activity is primarily regulated at the level of enzyme availability and post-translational modification. In E. coli, oxidative stress induced by peroxyl radicals leads to modification of specific amino acids, causing aggregation and inhibition of 6PGL. This suggests that under conditions of oxidative stress, 6PGL activity may be downregulated, potentially impacting PPP flux. In Arabidopsis, the presence of multiple peroxisomal isoforms suggests that regulation may occur through differential gene expression or protein targeting, although specific mechanisms are not fully elucidated. In parasites, the bifunctional G6PD-6PGL enzyme may be regulated by the availability of substrates or by feedback inhibition, but detailed regulatory pathways remain to be characterized [7,8]. Overall, regulation of 6PGL is an emerging area of research, with links to cellular redox status and metabolic demands.
6-phosphogluconolactonase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| PGLS (human) | Cancer metabolism | Knockout in cancer cell lines to assess proliferation and sensitivity to oxidative stress |
| G6PD-6PGL (P. falciparum) | Malaria | Enzyme inhibition assays and parasite growth inhibition |
| LdPGL (Leishmania donovani) | Leishmaniasis | Gene knockout in parasites to test infectivity |
| pgl (E. coli) | Oxidative stress response | Point mutations to mimic oxidative modifications |
| AtPGL (Arabidopsis) | Plant peroxisomal metabolism | Knockout lines to study redundancy and stress tolerance |
Cancer Metabolism and Imaging
Cancer cells often exhibit increased flux through the pentose phosphate pathway to support rapid proliferation and manage oxidative stress. 6PGL activity is a key component of this pathway, and its inhibition could disrupt cancer metabolism. Hyperpolarized δ-[1-13C]gluconolactone has been used to image 6PGL activity in brain tumors in vivo, demonstrating that this enzyme can serve as a metabolic imaging biomarker. This technique allows non-invasive assessment of PPP flux and could be used to monitor treatment response.
Parasitic Infections
In Leishmania donovani, 6PGL is crucial for growth and host infection, making it a potential drug target for leishmaniasis. Similarly, the bifunctional G6PD-6PGL enzyme in Plasmodium falciparum and Plasmodium vivax is essential for the parasite's metabolism and has been explored as a target for antimalarial drugs [7,8]. Inhibitors of 6PGL activity could selectively kill parasites by disrupting their PPP.
Oxidative Stress and Protein Aggregation
In E. coli, peroxyl radicals modify 6PGL, leading to aggregation and loss of activity. This suggests that under oxidative stress, 6PGL may be a target for damage, potentially contributing to metabolic dysfunction. In higher organisms, similar oxidative modifications could impair PPP flux and contribute to diseases associated with oxidative stress, such as neurodegeneration, although direct evidence is still limited.
From 6-phosphogluconolactonase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of 6PGL affect cancer cell proliferation? | PGLS knockout in cancer cell lines (e.g., HeLa, MCF-7) |
| Can 6PGL be targeted for antimalarial therapy? | G6PD-6PGL knockout or point mutation in P. falciparum |
| What is the role of 6PGL in oxidative stress? | pgl point mutations in E. coli to mimic oxidative damage |
| Is 6PGL essential for Leishmania infection? | LdPGL knockout in L. donovani |
| Do peroxisomal 6PGL isoforms have redundant functions? | Multiple AtPGL knockouts in Arabidopsis |
| Can 6PGL activity be imaged in vivo? | Overexpression of PGLS in tumor xenografts for hyperpolarized MRI |
How to Study the 6-phosphogluconolactonase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Enzymatic assay | 6PGL activity via coupled NADPH production | Kinetic characterization and inhibitor screening |
| Hyperpolarized MRI | 6PGL activity in vivo | Tumor imaging and metabolic flux |
| CRISPR knockout | Loss-of-function effects | Studying gene essentiality in cell lines and parasites [3,4] |
| CRISPR point mutation | Specific amino acid changes | Mimicking oxidative modifications or catalytic residues |
| Proteomics | Protein modifications and interactions | Identifying oxidative damage sites |
| RNA-seq | Gene expression changes | Assessing transcriptional regulation of PPP genes |
| Metabolomics | Metabolite levels | Measuring PPP flux and lactone accumulation |
| Inhibitor studies | Enzyme inhibition | Drug discovery for parasites and cancer [7,8] |
Enzymatic Activity Assays
6PGL activity can be measured spectrophotometrically by monitoring the hydrolysis of 6-phosphogluconolactone to 6-phosphogluconate, often coupled to NADP+ reduction via 6-phosphogluconate dehydrogenase. This assay is useful for characterizing enzyme kinetics and screening inhibitors. In E. coli, activity assays have been used to demonstrate inhibition by peroxyl radicals.
Hyperpolarized Magnetic Resonance Imaging
Hyperpolarized δ-[1-13C]gluconolactone is a substrate for 6PGL that can be detected by magnetic resonance spectroscopy and imaging. This technique has been used to image 6PGL activity in brain tumors in vivo, providing a non-invasive readout of PPP flux. It allows real-time assessment of enzyme activity in living organisms.
CRISPR-Cas9 Genome Editing
CRISPR-Cas9 can be used to generate knockout, point mutation, knock-in, or overexpression models for genes encoding 6PGL or its fused forms. For example, knockout of PGLS in human cell lines can reveal its role in metabolism and stress response. In parasites, CRISPR-mediated knockout of G6PD-6PGL can validate drug targets [7,8].
Proteomics and Oxidative Modification Analysis
Mass spectrometry-based proteomics can identify oxidative modifications on 6PGL, such as those induced by peroxyl radicals in E. coli. This approach helps map vulnerable residues and understand how oxidative stress regulates enzyme activity. It can also be used to study post-translational modifications in higher organisms.
How CRISPR Can Be Used to Study GO:0017057 6-phosphogluconolactonase activity
Knockout
CRISPR-Cas9 knockout of PGLS or its homologs can completely abolish 6PGL activity, allowing researchers to study its role in metabolism, growth, and stress response. For example, knockout of PGLS in cancer cell lines can reveal whether cells become dependent on alternative pathways for NADPH production. In Leishmania donovani, knockout of LdPGL reduces growth and infectivity, validating it as a drug target.
Point Mutation
Point mutations can be introduced into the active site or regulatory residues of 6PGL to dissect catalytic mechanism and regulation. For instance, mimicking oxidative modifications identified in E. coli (e.g., specific amino acid oxidations) can test their impact on enzyme activity and aggregation. Such models are valuable for understanding how oxidative stress affects PPP flux.
Knock-in
Knock-in of tagged 6PGL (e.g., GFP or FLAG) allows for localization and interaction studies. This can be used to track the enzyme in live cells and tissues, and to purify protein complexes for proteomic analysis. In Arabidopsis, knock-in of tagged peroxisomal 6PGL isoforms could help determine their subcellular localization and redundancy.
Overexpression
Overexpression of 6PGL can increase PPP flux and NADPH production, which may protect cells from oxidative stress or promote tumor growth. In cancer research, overexpression models can test whether elevated 6PGL activity contributes to metabolic reprogramming. In parasites, overexpression can be used to study drug resistance mechanisms.
How EDITGENE Supports 6-phosphogluconolactonase activity Research
Researchers studying 6-phosphogluconolactonase activity-related genes often need to determine whether a candidate gene is causally involved in metabolic regulation, disease progression, or drug response. EDITGENE provides a comprehensive suite of CRISPR-based services to accelerate this research, from gene knockout to precise point mutations and library screening.
Contact EDITGENE today to design your custom CRISPR model for 6-phosphogluconolactonase activity research.
Frequently Asked Questions About 6-phosphogluconolactonase activity
What is 6-phosphogluconolactonase activity?
6-phosphogluconolactonase activity (GO:0017057) is the enzymatic hydrolysis of 6-O-phosphono-D-glucono-1,5-lactone to 6-phospho-D-gluconate and H+, a key step in the pentose phosphate pathway.
What genes are involved in 6-phosphogluconolactonase activity?
Genes include PGLS in humans, pgl in E. coli, G6PD-6PGL in Plasmodium species, H6PD in mouse, and AtPGL isoforms in Arabidopsis [2,3,5,6,7,8].
Why is 6-phosphogluconolactonase important for metabolism?
It prevents the accumulation of a reactive lactone intermediate and maintains flux through the pentose phosphate pathway, which supplies NADPH and ribose-5-phosphate.
How is 6-phosphogluconolactonase activity measured?
It can be measured by coupled enzymatic assays monitoring NADPH production, or by hyperpolarized MRI using δ-[1-13C]gluconolactone [1,3].
What diseases are associated with 6-phosphogluconolactonase?
It is linked to cancer metabolism, parasitic infections such as malaria and leishmaniasis, and oxidative stress-related conditions [1,4,7,8].
Can 6-phosphogluconolactonase be a drug target?
Yes, in parasites like Plasmodium and Leishmania, 6PGL is essential for survival and has been explored as a drug target [4,7,8].
How does oxidative stress affect 6-phosphogluconolactonase?
In E. coli, peroxyl radicals modify specific amino acids, causing aggregation and inhibition of 6PGL activity.
What is the role of 6-phosphogluconolactonase in cancer?
Cancer cells often upregulate the pentose phosphate pathway; 6PGL activity supports NADPH production and can be imaged in tumors [1,3].
Are there fused forms of 6-phosphogluconolactonase?
Yes, in Plasmodium species and mouse, 6PGL is fused to glucose-6-phosphate dehydrogenase, forming a bifunctional enzyme [5,7,8].
How can CRISPR be used to study 6-phosphogluconolactonase?
CRISPR can create knockout, point mutation, knock-in, or overexpression models to dissect the gene's function in metabolism and disease [3,4].
Conclusion
6-phosphogluconolactonase activity (GO:0017057) is a fundamental enzymatic function in the pentose phosphate pathway, with critical roles in metabolism, oxidative stress response, and disease. Its importance spans from bacterial stress responses to parasite survival and cancer metabolism. Advances in imaging and CRISPR-based models are providing new insights into its regulation and potential as a therapeutic target. Continued research on 6PGL will likely uncover further links to human health and disease.
References
- 1. Batsios G et al.. 2021. Imaging 6-Phosphogluconolactonase Activity in Brain Tumors In Vivo Using Hyperpolarized δ-[1-(13)C]gluconolactone.. Front Oncol 11:589570 PMID: 33937017
- 2. Reyes JS et al.. 2023. Peroxyl radicals modify 6-phosphogluconolactonase from Escherichia coli via oxidation of specific amino acids and aggregation which inhibits enzyme activity.. Free Radic Biol Med 204:118-127 PMID: 37119864
- 3. Phégnon L et al.. 2024. 6-Phosphogluconolactonase is critical for the efficient functioning of the pentose phosphate pathway.. FEBS J 291(20):4459-4472 PMID: 38982839
- 4. Paul A et al.. 2023. Leishmania donovani 6-phosphogluconolactonase: Crucial for growth and host infection?. Microb Pathog 178:106082 PMID: 36958644
- 5. Clarke JL et al.. 2003. Murine hexose-6-phosphate dehydrogenase: a bifunctional enzyme with broad substrate specificity and 6-phosphogluconolactonase activity.. Arch Biochem Biophys 415(2):229-34 PMID: 12831846
- 6. Lansing H et al.. 2020. Analysis of potential redundancy among Arabidopsis 6-phosphogluconolactonase isoforms in peroxisomes.. J Exp Bot 71(3):823-836 PMID: 31641750
- 7. Allen SM et al.. 2015. Plasmodium falciparum glucose-6-phosphate dehydrogenase 6-phosphogluconolactonase is a potential drug target.. FEBS J 282(19):3808-23 PMID: 26198663
- 8. Haeussler K et al.. 2019. Glucose 6-phosphate dehydrogenase 6-phosphogluconolactonase: characterization of the Plasmodium vivax enzyme and inhibitor studies.. Malar J 18(1):22 PMID: 30683097
