scholarly journals Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from Toxoplasma gondii

2021 ◽  
Vol 22 (15) ◽  
pp. 7830
Author(s):  
Rajib Acharjee ◽  
Keith K. Talaam ◽  
Endah D. Hartuti ◽  
Yuichi Matsuo ◽  
Takaya Sakura ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Drug Target ◽  
Tricarboxylic Acid Cycle ◽  
Expression System ◽  
Protozoan Parasite ◽  
New Drugs ◽  
Quinone Oxidoreductase ◽  
Steady State Kinetics ◽  
Transport Chain ◽  
Effective Drugs

Toxoplasma gondii is a protozoan parasite that causes toxoplasmosis and infects almost one-third of the global human population. A lack of effective drugs and vaccines and the emergence of drug resistant parasites highlight the need for the development of new drugs. The mitochondrial electron transport chain (ETC) is an essential pathway for energy metabolism and the survival of T. gondii. In apicomplexan parasites, malate:quinone oxidoreductase (MQO) is a monotopic membrane protein belonging to the ETC and a key member of the tricarboxylic acid cycle, and has recently been suggested to play a role in the fumarate cycle, which is required for the cytosolic purine salvage pathway. In T. gondii, a putative MQO (TgMQO) is expressed in tachyzoite and bradyzoite stages and is considered to be a potential drug target since its orthologue is not conserved in mammalian hosts. As a first step towards the evaluation of TgMQO as a drug target candidate, in this study, we developed a new expression system for TgMQO in FN102(DE3)TAO, a strain deficient in respiratory cytochromes and dependent on an alternative oxidase. This system allowed, for the first time, the expression and purification of a mitochondrial MQO family enzyme, which was used for steady-state kinetics and substrate specificity analyses. Ferulenol, the only known MQO inhibitor, also inhibited TgMQO at IC50 of 0.822 μM, and displayed different inhibition kinetics compared to Plasmodium falciparum MQO. Furthermore, our analysis indicated the presence of a third binding site for ferulenol that is distinct from the ubiquinone and malate sites.

scholarly journals Identification of Plasmodium falciparum Mitochondrial Malate: Quinone Oxidoreductase Inhibitors from the Pathogen Box

Genes ◽  
2019 ◽  
Vol 10 (6) ◽  
pp. 471 ◽  
Author(s):  
Xinying Wang ◽  
Yukiko Miyazaki ◽  
Daniel Ken Inaoka ◽  
Endah Dwi Hartuti ◽  
Yoh-Ichi Watanabe ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Antimalarial Drug ◽  
Drug Target ◽  
Drug Targets ◽  
Tricarboxylic Acid Cycle ◽  
Drug Resistant ◽  
Quinone Oxidoreductase ◽  
Mitochondrial Inner Membrane ◽  
Transport Chain ◽  
Urgent Task

Malaria is one of the three major global health threats. Drug development for malaria, especially for its most dangerous form caused by Plasmodium falciparum, remains an urgent task due to the emerging drug-resistant parasites. Exploration of novel antimalarial drug targets identified a trifunctional enzyme, malate quinone oxidoreductase (MQO), located in the mitochondrial inner membrane of P. falciparum (PfMQO). PfMQO is involved in the pathways of mitochondrial electron transport chain, tricarboxylic acid cycle, and fumarate cycle. Recent studies have shown that MQO is essential for P. falciparum survival in asexual stage and for the development of experiment cerebral malaria in the murine parasite P. berghei, providing genetic validation of MQO as a drug target. However, chemical validation of MQO, as a target, remains unexplored. In this study, we used active recombinant protein rPfMQO overexpressed in bacterial membrane fractions to screen a total of 400 compounds from the Pathogen Box, released by Medicines for Malaria Venture. The screening identified seven hit compounds targeting rPfMQO with an IC50 of under 5 μM. We tested the activity of hit compounds against the growth of 3D7 wildtype strain of P. falciparum, among which four compounds showed an IC50 from low to sub-micromolar concentrations, suggesting that PfMQO is indeed a potential antimalarial drug target.

scholarly journals The ubiquinone synthesis pathway is a promising drug target for Chagas disease

PLoS ONE ◽  
2021 ◽  
Vol 16 (2) ◽  
pp. e0243855
Author(s):  
Takeshi Nara ◽  
Yukari Nakagawa ◽  
Keiko Tsuganezawa ◽  
Hitomi Yuki ◽  
Katsuhiko Sekimata ◽  
...  
Keyword(s):  
Chagas Disease ◽  
Latin American ◽  
Drug Target ◽  
Culture Medium ◽  
Protozoan Parasite ◽  
Redox Balance ◽  
New Drugs ◽  
Key Enzyme ◽  
2D And 3D ◽  
Similarity Searches

Chagas disease is caused by infection with the protozoan parasite Trypanosoma cruzi (T. cruzi). It was originally a Latin American endemic health problem, but now is expanding worldwide as a result of increasing migration. The currently available drugs for Chagas disease, benznidazole and nifurtimox, provoke severe adverse effects, and thus the development of new drugs is urgently required. Ubiquinone (UQ) is essential for respiratory chain and redox balance in trypanosomatid protozoans, therefore we aimed to provide evidence that inhibitors of the UQ biosynthesis have trypanocidal activities. In this study, inhibitors of the human COQ7, a key enzyme of the UQ synthesis, were tested for their trypanocidal activities because they were expected to cross-react and inhibit trypanosomal COQ7 due to their genetic homology. We show the trypanocidal activity of a newly found human COQ7 inhibitor, an oxazinoquinoline derivative. The structurally similar compounds were selected from the commercially available compounds by 2D and 3D ligand-based similarity searches. Among 38 compounds selected, 12 compounds with the oxazinoquinoline structure inhibited significantly the growth of epimastigotes of T. cruzi. The most effective 3 compounds also showed the significant antitrypanosomal activity against the mammalian stage of T. cruzi at lower concentrations than benznidazole, a commonly used drug today. We found that epimastigotes treated with the inhibitor contained reduced levels of UQ9. Further, the growth of epimastigotes treated with the inhibitors was partially rescued by UQ10 supplementation to the culture medium. These results suggest that the antitrypanosomal mechanism of the oxazinoquinoline derivatives results from inhibition of the trypanosomal UQ synthesis leading to a shortage of the UQ pool. Our data indicate that the UQ synthesis pathway of T. cruzi is a promising drug target for Chagas disease.

scholarly journals TgATAT-Mediated α-Tubulin Acetylation Is Required for Division of the Protozoan Parasite Toxoplasma gondii

mSphere ◽  
2016 ◽  
Vol 1 (1) ◽  
Author(s):  
Joseph M. Varberg ◽  
Leah R. Padgett ◽  
Gustavo Arrizabalaga ◽  
William J. Sullivan
Keyword(s):  
Toxoplasma Gondii ◽  
Drug Target ◽  
Drug Targets ◽  
Protozoan Parasite ◽  
World Population ◽  
Content Type ◽  
Tubulin Acetylation ◽  
Parasite Replication ◽  
New Treatments

ABSTRACT Toxoplasma gondii is an opportunistic parasite that infects at least one-third of the world population. New treatments for the disease (toxoplasmosis) are needed since current drugs are toxic to patients. Microtubules are essential cellular structures built from tubulin that show promise as antimicrobial drug targets. Microtubules can be regulated by chemical modification, such as acetylation on lysine 40 (K40). To determine the role of K40 acetylation in Toxoplasma and whether it is a liability to the parasite, we performed mutational analyses of the α-tubulin gene. Our results indicate that parasites cannot survive without K40 acetylation unless microtubules are stabilized with a secondary mutation. Additionally, we identified the parasite enzyme that acetylates α-tubulin (TgATAT). Genetic disruption of TgATAT caused severe defects in parasite replication, further highlighting the importance of α-tubulin K40 acetylation in Toxoplasma and its promise as a potential new drug target. Toxoplasma gondii is a widespread protozoan parasite that causes potentially life-threatening opportunistic disease. New inhibitors of parasite replication are urgently needed, as the current antifolate treatment is also toxic to patients. Microtubules are essential cytoskeletal components that have been selectively targeted in microbial pathogens; further study of tubulin in Toxoplasma may reveal novel therapeutic opportunities. It has been noted that α-tubulin acetylation at lysine 40 (K40) is enriched during daughter parasite formation, but the impact of this modification on Toxoplasma division and the enzyme mediating its delivery have not been identified. We performed mutational analyses to provide evidence that K40 acetylation stabilizes Toxoplasma microtubules and is required for parasite replication. We also show that an unusual Toxoplasma homologue of α-tubulin acetyltransferase (TgATAT) is expressed in a cell cycle-regulated manner and that its expression peaks during division. Disruption of TgATAT with CRISPR/Cas9 ablates K40 acetylation and induces replication defects; parasites appear to initiate mitosis yet exhibit incomplete or improper nuclear division. Together, these findings establish the importance of tubulin acetylation, exposing a new vulnerability in Toxoplasma that could be pharmacologically targeted. IMPORTANCE Toxoplasma gondii is an opportunistic parasite that infects at least one-third of the world population. New treatments for the disease (toxoplasmosis) are needed since current drugs are toxic to patients. Microtubules are essential cellular structures built from tubulin that show promise as antimicrobial drug targets. Microtubules can be regulated by chemical modification, such as acetylation on lysine 40 (K40). To determine the role of K40 acetylation in Toxoplasma and whether it is a liability to the parasite, we performed mutational analyses of the α-tubulin gene. Our results indicate that parasites cannot survive without K40 acetylation unless microtubules are stabilized with a secondary mutation. Additionally, we identified the parasite enzyme that acetylates α-tubulin (TgATAT). Genetic disruption of TgATAT caused severe defects in parasite replication, further highlighting the importance of α-tubulin K40 acetylation in Toxoplasma and its promise as a potential new drug target.

scholarly journals Divergent features of the coenzyme Q:cytochrome c oxidoreductase complex in Toxoplasma gondii parasites

2021 ◽  
Vol 17 (2) ◽  
pp. e1009211
Author(s):  
Jenni A. Hayward ◽  
Esther Rajendran ◽  
Soraya M. Zwahlen ◽  
Pierre Faou ◽  
Giel G. van Dooren
Keyword(s):  
Toxoplasma Gondii ◽  
Drug Target ◽  
Protein Composition ◽  
Complex Iii ◽  
Mitochondrial Electron Transport Chain ◽  
Eukaryotic Evolution ◽  
Mitochondrial Oxygen Consumption ◽  
Apicomplexan Parasites ◽  
Transport Chain ◽  
Important Drug

The mitochondrion is critical for the survival of apicomplexan parasites. Several major anti-parasitic drugs, such as atovaquone and endochin-like quinolones, act through inhibition of the mitochondrial electron transport chain at the coenzyme Q:cytochrome c oxidoreductase complex (Complex III). Despite being an important drug target, the protein composition of Complex III of apicomplexan parasites has not been elucidated. Here, we undertake a mass spectrometry-based proteomic analysis of Complex III in the apicomplexan Toxoplasma gondii. Along with canonical subunits that are conserved across eukaryotic evolution, we identify several novel or highly divergent Complex III components that are conserved within the apicomplexan lineage. We demonstrate that one such subunit, which we term TgQCR11, is critical for parasite proliferation, mitochondrial oxygen consumption and Complex III activity, and establish that loss of this protein leads to defects in Complex III integrity. We conclude that the protein composition of Complex III in apicomplexans differs from that of the mammalian hosts that these parasites infect.

scholarly journals Divergent features of the coenzyme Q:cytochrome c oxidoreductase complex in Toxoplasma gondii parasites

2020 ◽  
Author(s):  
Jenni A. Hayward ◽  
Esther Rajendran ◽  
Soraya M. Zwahlen ◽  
Pierre Faou ◽  
Giel G. van Dooren
Keyword(s):  
Electron Transport ◽  
Toxoplasma Gondii ◽  
Electron Transport Chain ◽  
Drug Target ◽  
Complex Function ◽  
Protein Composition ◽  
Complex Iii ◽  
Mitochondrial Electron Transport Chain ◽  
Apicomplexan Parasites ◽  
Transport Chain

AbstractThe mitochondrion is critical for the survival of apicomplexan parasites. Several major anti-parasitic drugs, such as atovaquone and endochin-like quinolones, act through inhibition of the mitochondrial electron transport chain at the coenzyme Q:cytochrome c oxidoreductase complex (Complex III). Despite being an important drug target, the protein composition of Complex III of apicomplexan parasites has not been elucidated. Here, we undertake a mass spectrometry-based proteomic analysis of Complex III in the apicomplexan Toxoplasma gondii. Along with canonical subunits that are conserved across eukaryotic evolution, we identify several novel or highly divergent Complex III components that are conserved within the apicomplexan lineage. We demonstrate that one such subunit, which we term TgQCR11, is critical for parasite proliferation, mitochondrial oxygen consumption and Complex III activity, and establish that loss of this protein leads to defects in Complex III integrity. We conclude that the protein composition of Complex III in apicomplexans differs from that of the mammalian hosts that these parasites infect.Author summaryApicomplexan parasites cause numerous diseases in humans and animals, including malaria (Plasmodium species) and toxoplasmosis (Toxoplasma gondii). The coenzyme Q:cytochrome c oxidoreductase protein complex (Complex III) performs a central role in the mitochondrial electron transport chain of many eukaryotes. Despite being the target of several major anti-apicomplexan drugs, the protein composition of Complex III in apicomplexans was previously unknown. Our work identifies novel proteins in Complex III of apicomplexans, one of which is critical for complex function and integrity. Our study highlights divergent features of Complex III in apicomplexans, and provides a broader understanding of Complex III evolution in eukaryotes. Our study also provides important insights into what sets this major drug target apart from the equivalent complex in host species.

scholarly journals Discovery of New Inhibitors of Toxoplasma gondii via the Pathogen Box

2017 ◽  
Vol 62 (2) ◽  
Author(s):  
Jérémy Spalenka ◽  
Sandie Escotte-Binet ◽  
Ali Bakiri ◽  
Jane Hubert ◽  
Jean-Hugues Renault ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Resistance Mechanism ◽  
Protozoan Parasite ◽  
Vero Cells ◽  
Selectivity Index ◽  
New Drugs ◽  
Enzyme Linked Immunosorbent Assay ◽  
Content Type ◽  
Drug Intolerance

ABSTRACT Toxoplasma gondii is a cosmopolitan protozoan parasite which affects approximately 30% of the population worldwide. The drugs currently used against toxoplasmosis are few in number and show several limitations, such as drug intolerance, poor bioavailability, or drug resistance mechanism developed by the parasite. Thus, it is important to find new compounds able to inhibit parasite invasion or proliferation. In this study, the 400 compounds of the open-access Pathogen Box, provided by the Medicines for Malaria Venture (MMV) foundation, were screened for their anti-Toxoplasma gondii activity. A preliminary in vitro screening performed over 72 h by an enzyme-linked immunosorbent assay (ELISA) revealed 15 interesting compounds that were effective against T. gondii at 1 μM. Their cytotoxicity was estimated on Vero cells, and their 50% inhibitory concentrations (IC50) were further calculated. As a result, eight anti-Toxoplasma gondii compounds with an IC50 of less than 2 μM and a selectivity index (SI) value of greater than 4 were identified. The most active was MMV675968, showing an IC50 of 0.02 μM and a selectivity index value equal to 275. Two other compounds, MMV689480 and MMV687807, also showed a good activity against T. gondii, with IC50s of 0.10 μM (SI of 86.6) and 0.15 μM (SI of 11.3), respectively. Structure-activity relationships for the eight selected compounds also were discussed on the basis of fingerprinting similarity measurements using the Tanimoto method. The anti-Toxoplasma gondii compounds highlighted here represent potential candidates for the development of new drugs that could be used against toxoplasmosis.

scholarly journals Toxoplasmicidal in vitro effect of dehydroepiandrosterone on Toxoplasma gondii extracellular tachizoytes

2020 ◽  
Author(s):  
Saé Muñiz-Hernández ◽  
Carmen T. Gómez de León ◽  
Angélica Luna Nophal ◽  
Lenin Domínguez-Ramírez ◽  
Olga-Araceli Patrón-Soberano ◽  
...  
Keyword(s):  
Toxoplasma Gondii ◽  
Adverse Effects ◽  
Molecular Level ◽  
Protozoan Parasite ◽  
New Drugs ◽  
Limited Effectiveness ◽  
Vitro Effect ◽  
First Time ◽  
In Vitro Treatment

AbstractToxoplasmosis is a zoonotic disease caused by the apicomplexa protozoan parasite Toxoplasma gondii. This disease is a health burden, mainly in pregnant women and immunocompromised individuals, in whom they can cause death. Despite advances in the medical area, nowadays there are no new drugs to treat toxoplasmosis. The standard therapy to toxoplasmosis has not had progress for last seven decades; it is a combination of sulfadiazine-pyrimethamine (S-P); which is co-administered with folic acid due to the adverse effects of the drug. Several studies have shown that the conventional treatment has limited effectiveness and severe adverse effects. Thus, the search of better treatments with greater efficacy and without the adverse effects becomes relevant. In the current work we demonstrate for the first time the parasiticidal effect of dehydroepiandrosterone (DHEA), a steroid hormone produced by many mammals, on extracellular tachyzoites (the infective stage of T. gondii). In vitro treatment with DHEA reduces the viability of extracellular tachyzoites, and both the active and passive invasion processes. The ultrastructural analysis of treated parasites showed that DHEA alters the cytoskeleton structures, leading in the lost of the organelle structure and organization, as well as, the lost of the cellular shape. On a molecular level, we observed an important reduction of the expression of several proteins that are essential for the motility and virulence of parasites when they were exposed to DHEA. These results suggest that DHEA could be used as an alternative treatment against toxoplasmosis.

A Drug Target Interaction Prediction Based on LINE-RF Learning

2020 ◽  
Vol 15 (7) ◽  
pp. 750-757
Author(s):  
Jihong Wang ◽  
Yue Shi ◽  
Xiaodan Wang ◽  
Huiyou Chang
Keyword(s):  
Network Topology ◽  
Drug Target ◽  
Large Scale ◽  
Representation Learning ◽  
New Drugs ◽  
Combination Method ◽  
Learning Methods ◽  
Network Representation ◽  
On Line ◽  
Clinical Experiments

Background: At present, using computer methods to predict drug-target interactions (DTIs) is a very important step in the discovery of new drugs and drug relocation processes. The potential DTIs identified by machine learning methods can provide guidance in biochemical or clinical experiments. Objective: The goal of this article is to combine the latest network representation learning methods for drug-target prediction research, improve model prediction capabilities, and promote new drug development. Methods: We use large-scale information network embedding (LINE) method to extract network topology features of drugs, targets, diseases, etc., integrate features obtained from heterogeneous networks, construct binary classification samples, and use random forest (RF) method to predict DTIs. Results: The experiments in this paper compare the common classifiers of RF, LR, and SVM, as well as the typical network representation learning methods of LINE, Node2Vec, and DeepWalk. It can be seen that the combined method LINE-RF achieves the best results, reaching an AUC of 0.9349 and an AUPR of 0.9016. Conclusion: The learning method based on LINE network can effectively learn drugs, targets, diseases and other hidden features from the network topology. The combination of features learned through multiple networks can enhance the expression ability. RF is an effective method of supervised learning. Therefore, the Line-RF combination method is a widely applicable method.

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