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Ornithine and proline metabolism. ODC, ornithine decarboxylase; P5CR, P5C reductase; P5CS, P5C synthase. Broken lines represent reactions that have not been demonstrated in P. falciparum.
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Ornithine δ-aminotransferase (OAT) of the malaria parasite Plasmodium falciparum catalyzes the reversible conversion of ornithine into glutamate-5-semialdehyde and glutamate and is-in contrast to its human counterpart-activated by thioredoxin (Trx) by a factor of 10. Trx, glutaredoxin, and plasmoredoxin are redox-active proteins that play a crucial...
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... the degradation of L-ornithine, OAT catalyzes the interconversion of ornithine and α-ketoglutarate (α-KG) into glutamate-5-semialdehyde, which spontaneously cyclizes to pyrroline-5-carboxylate (P5C) and glutamate. 15,16 Thus, it is required for the production of proline and glutamate from ornithine and controls the catabolism of excess ornithine (Fig. 1). In addition, plasmodial OAT is capable of synthesizing ornithine from glutamine via glutamate (an inferred conversion into glutamate-5-phosphate and glutamate-5-semialdehyde). A previous study using radiolabeled glutamine, however, demonstrated that production of ornithine by PfOAT contributes only marginally to the overall ornithine ...
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... to prevent the toxic accumulation of ornithine. 16 In addition, PfOAT directly influences the synthesis of polyamines by regulating ornithine levels 19 and is involved in the regulation of mitotic cell division. 20 PfOAT is, with 16 cysteine residues, highly cysteine rich; only three of these cysteines are conserved in the OAT of most organisms (Fig. S1). Human OAT (hOAT) is located in the mitochondrial matrix, 21,22 whereas plasmodial OAT, similar to OAT from Saccharomyces cerevisiae, 23 lacks the mitochondrial targeting sequence and is therefore supposed to be located in the cytosol. hOAT has been extensively studied both biochemically and genetically mainly because of its ...
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... data suggest that Trx reduces a disulfide between Cys154 and Cys163 of one subunit. Cys154 and Cys163 are highly conserved in Plasmodium and mediate activity regulation by Trx, but cannot be found in other organisms (Fig. S1). The interaction remains to be studied in detail by cocrystallization experiments of PfOAT and ...
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... can be synthesized via two pathways that share P5C as an intermediate (Fig. 1). 50 The malaria parasite codes for a putative P5C reductase, but no gene with similarity to P5C synthase, glutamate 5-kinase, or glutamate 5-semialdehyde dehydrogenase can be found in the genome of Plasmodium. This suggests that ornithine degradation via PfOAT might be the only pathway of proline production in Plasmodium species. ...
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... The protein S-glutathionylation reaction can affect the function of modified proteins ( Figure 6C). A notable example is P. falciparum ornithine aminotransferase (PfOAT, PF3D7_0608800), which directly affects polyamine synthesis by regulating ornithine levels and is involved in the regulation of glucose metabolism and cell proliferation [40,41]. The active site Cys residues 154 and 163 of PfOAT can be modified by S-glutathionylation to form disulfide bond, resulting in impaired substrate binding and hence catalytic activity [42]. ...
Malaria continues to pose a serious global health threat, and artemisinin remains the core drug for global malaria control. However, the situation of malaria resistance has become increasingly severe due to the emergence and spread of artemisinin resistance. In recent years, significant progress has been made in understanding the mechanism of action (MoA) of artemisinin. Prior research on the MoA of artemisinin mainly focused on covalently bound targets that are alkylated by artemisinin‐free radicals. However, less attention has been given to the reversible noncovalent binding targets, and there is a paucity of information regarding artemisinin targets at different life cycle stages of the parasite. In this study, we identified the protein targets of artemisinin at different stages of the parasite's intraerythrocytic developmental cycle using a photoaffinity probe. Our findings demonstrate that artemisinin interacts with parasite proteins in vivo through both covalent and noncovalent modes. Extensive mechanistic studies were then conducted by integrating target validation, phenotypic studies, and untargeted metabolomics. The results suggest that protein synthesis, glycolysis, and oxidative homeostasis are critically involved in the antimalarial activities of artemisinin. In summary, this study provides fresh insights into the mechanisms underlying artemisinin's antimalarial effects and its protein targets.
... The binding pocket location was mapped according to the information reported by the previous study. 14 ...
Four novel 4-aminoquinoline-isoindolin-1-one hybrids 5a-d have been designed as antiplasmodial and antioxidant agents by combining the 4-aminoquinoline unit and isoindolin-1-one moiety using the flexible alkyl linker. The hybrids were synthesized in two steps via aromatic nucleophilic substitution and nucleophilic addition reactions and were obtained with yields up to 83%. The evaluation of their anti-plasmodial activity was conducted against Plasmodium falciparum 3D7, whereas the antioxidant activity was assessed using the DPPH method. Among the four hybrids, the hybrid of 3-benzyl-2-(6-((7-chloroquinolin-4-yl)amino)hexyl)-3-hydroxyisoindolin-1-one 5c demonstrated excellent antiplasmodial (IC50 of 0.01 µg/mL) and antioxidant (IC50 of 46.58 µg/mL) activities. The molecular docking study was conducted using PfOAT as the protein target. The docking results were in line with the in vitro antiplasmodial assay, where hybrid 5c displayed the lowest binding energy of -8.43 kcal/mol
... P5C is a spontaneously rearranged product of glutamate-5semialdehyde that forms a colored product upon reaction with ninhydrin, which can be monitored by absorbance at 512 nm, reporting on OAT enzymatic activity. 69,70 Nitrosated OAT was obtained by treatment of wild-type OAT with GSNO. The activity of the nitrosated wild-type OAT was slightly lower, 64.5%, compared to the non-nitrosated control, although the changes were not significant (p > 0.05) ( Figure 5F). ...
... The purified OAT enzyme activity was assayed by monitoring the production of glutamate-5-semialdehyde via absorbance at 512 nm, as previously described. 69,70 OAT WT, C150S, or C150A [100 μL of 0.25 μg/mL in HEN buffer (or 5.6 μM)] were preincubated with 1 mM GSNO (in HEN) for 1 h at 37°C ...
... Dimeric, tetrameric, and hexameric states have been described for monofunctional P5CDH (Srivastava et al., 2012;Luo et al., 2013;Pemberton and Tanner, 2013;Korasick et al., 2019). Monomeric, dimeric, and tetrameric structures have also been reported for OAT (Sekhar et al., 2007;Jortzik et al., 2010;Stránská et al., 2010;Montioli et al., 2017). ...
Proline dehydrogenase (ProDH) and pyrroline-5-carboxylate (P5C) dehydrogenase (P5CDH) catalyze the oxidation of proline into glutamate via the intermediates P5C and glutamate-semialdehyde (GSA), which spontaneously interconvert. P5C and GSA are also intermediates in the production of glutamate from ornithine and α-ketoglutarate catalyzed by ornithine δ-aminotransferase (OAT). ProDH and P5CDH form a fused bifunctional PutA enzyme in Gram-negative bacteria and are associated in a bifunctional substrate channelling complex in Thermus thermophilus, but the physical proximity of ProDH and P5CDH in eukaryotes has not been described. Here we report evidence of physical proximity and interactions between Arabidopsis ProDH, P5CDH and OAT in the mitochondria of plants during dark-induced leaf senescence when all three enzymes are expressed. Pairwise interactions and localization of the three enzymes were investigated using bimolecular fluorescence complementation (BiFC) with confocal microscopy in tobacco and sub-mitochondrial fractionation in Arabidopsis. Evidence for a complex composed of ProDH, P5CDH, and OAT was revealed by co-migration of the proteins in native conditions upon gel electrophoresis. Co-immunoprecipitation coupled with mass spectrometry analysis confirmed the presence of the P5C metabolism complex in Arabidopsis. Pull-down assays further demonstrated a direct interaction between ProDH1 and P5CDH. P5C metabolism complexes may channel P5C among the constituent enzymes and directly provide electrons to the respiratory electron chain via ProDH.
... Several enzymes involved in the key metabolic pathways of the parasite were revealed by GO analysis, including ornithine aminotransferase (OAT), pyruvate kinase (PyrK), L-lactate dehydrogenase (LDH), spermidine synthase (SpdSyn), S-adenosylmethionine synthetase (SAMS) and P. falciparum hexose transporter 1 (PFHT1). 190,194,195,204,[218][219][220] The protein data from Research Collaboratory for Structural Bioinformatics PDB (RCSB PDB: https://www.rcsb.org/) and worldwide Protein Data Bank (wwPDB: http://www.wwpdb.org/) ...
Covering 1972 to 2021Malaria remains a significant public health problem in some regions of the world. The great efforts to control malaria have been severely compromised due to the widespread resistance of Plasmodium falciparum to nearly all frontline drugs. Pursuit of novel molecules from the sea will potentially result in new interventions against malaria, which are urgently needed to combat the increase of resistance. Focusing on the strategy of the "Blue Drug Bank", the molecules highlighted here can serve as an inspiration for future medicinal chemistry campaigns. This review covers the developments in the field of antimalarial marine lead compounds reported between 1972 and July 2021, and offers a comprehensive overview on their progresses and potentials. We selected 60 representative potential candidate molecules from 361 marine natural products, and highlighted their structure-activity relationships, molecular mechanisms of targets, and drug-like properties in order to assess their full potential to be developed. We summarized 107 clinically proven or potential antimalarial targets and their subcellular locations in the relevant target proteins, which linked the molecules to the target proteins at the subcellular level. Hence, it could be expected that natural products targeting different mechanisms may prove to be an effective strategy in antimalarial drug research and development in the future.
... PfOAT enzymatic activity by reducing a disulfide in a substrate-binding loop, thereby increasing the affinity of PfOAT toward its substrate and modulating polyamine synthesis and ornithine homeostasis in response to cellular redox state (Jortzik et al. 2010). The structural study of PfTrx1 by NMR spectroscopy illustrates that the disulfide formation leads to a change in PfTrx1 which may be essential (Munte et al. 2005). ...
... In a study, the Plrx increased transcription of Trx, TrxR, and Gr while Grx had no change (Buchholz et al. 2008). Another study reported the identification of 21 proteins, which interact with PfPlrx (Sturm et al. 2009b) enhancing the activity of Pf OAT but less efficiently than PfTrx1 (Jortzik et al. 2010). One common function shared by Pfolrx, PfTrx1, and PfGrx is the catalysis of deglutathionylation reaction of the parasite Plasmodium falciparum protein (Kehr et al. 2011). ...
The malaria-causing parasite Plasmodium falciparum is a severe threat to human health across the globe. This parasite alone causes the highest morbidity and mortality than any other species of Plasmodium. The parasites dynamically multiply in the erythrocytes of the vertebrate hosts, a large number of reactive oxygen species that damage biological macromolecules are produced in the cell during parasite growth. To relieve this intense oxidative stress, the parasite employs an NADPH-dependent thioredoxin and glutathione system that acts as an antioxidant and maintains redox status in the parasite. The mutual interaction of both redox proteins is involved in various biological functions and the survival of the erythrocytic stage of the parasite. Since the Plasmodium species is deficient in catalase and classical glutathione peroxidase, so their redox balance relies on a complex set of five peroxiredoxins, differentially positioned in the cytosol, mitochondria, apicoplast, and nucleus with partly overlapping substrate preferences. Moreover, Plasmodium falciparum possesses a set of members belonging to the thioredoxin superfamily, such as three thioredoxins, two thioredoxin-like proteins, one dithiol, three monocysteine glutaredoxins, and one redox-active plasmoredoxin with largely redundant functions. This review paper aims to discuss and encapsulate the biological function and current knowledge of the functional redox network of Plasmodium falciparum.
... Ornithine, via the actions of the ornithine aminotransferase (OAT) and pyrroline-5-carboxylate reductase enzymes, is then converted into proline. The P. falciparum, PfOAT was expressed in E. coli and was investigated for its activity [57,58]. PfOAT was shown to be bidirectional, functioning predominantly in the ornithine to proline direction [57]. ...
Obligate intracellular pathogens have coevolved with their host, leading to clever strategies to access nutrients, to combat the host’s immune response, and to establish a safe niche for intracellular replication. The host, on the other hand, has also developed ways to restrict the replication of invaders by limiting access to nutrients required for pathogen survival. In this review, we describe the recent advancements in both computational methods and high-throughput –omics techniques that have been used to study and interrogate metabolic functions in the context of intracellular parasitism. Specifically, we cover the current knowledge on the presence of amino acid biosynthesis and uptake within the Apicomplexa phylum, focusing on human-infecting pathogens: Toxoplasma gondii and Plasmodium falciparum. Given the complex multi-host lifecycle of these pathogens, we hypothesize that amino acids are made, rather than acquired, depending on the host niche. We summarize the stage specificities of enzymes revealed through transcriptomics data, the relevance of amino acids for parasite pathogenesis in vivo, and the role of their transporters. Targeting one or more of these pathways may lead to a deeper understanding of the specific contributions of biosynthesis versus acquisition of amino acids and to design better intervention strategies against the apicomplexan parasites.
... e in silico method being used is molecular docking as supplemented in Supplementary Materials (S1). e seven compounds (ligands) are precocene I, precocene II, VMDC, beta-sitosterol, stigmasterol, polymethoxyflavone, and pyrrolizidine, whereas the seven proteins are human tyrosinase (5m8s) [81], Plasmodium falciparum ornithine delta-aminotransferase (3lg0) [82], Mus musculus cyclooxygenase-2 (4cox) [83], E. coli 4-diphosphodicytidyl-2-C-methylerythritol synthase (1i52) [84], glycoside hydrolase family 97 from Bacteroides thetaiotaomicron (2zq0) [85], human p53 (4ibu) [86], and human MMP-9 (1l6j) [87]. ...
Goat weed (Ageratum conyzoides L.), or bandotan in Indonesia, is an herbaceous plant that broadly grows up in both subtropical as well as tropical areas. is herb contains many phytoconstituents which have many benefits in different aspects. e essential oil contains phytochemicals such as phenol, phenolic ester, and coumarin, whereas many compounds can been identified in the whole part such as terpenoid, steroid, chromene, pyrrolizidine alkaloid, and flavonoid. Empirically, this herb has been used as an antihemorrhagic, antiseptic, antileprosy, and wound-healing agent. is article reviews the potency of the herb in medication according to the chemical substances being deposited, which are collected from numerous studies, followed by its in silico bioactivity prediction as well as its pharmaceutical dosage form formulation.
... The crystal structure of TgOAT in complex with pyrodoxal-5 ′phosphate (PLP) was determined at 1.2 Å resolution (PDB 4nog; Figure 6A and Table 2) and contains two protein chains in the asymmetric unit (∼0.3 Å r.m.s.d. over 422 Cα atoms) that form a well-studied functional dimer with BSA of ∼5,770 Å 2 (Shah et al., 1997;Shen et al., 1998;Markova et al., 2005;Vedadi et al., 2007;Jortzik et al., 2010) (Figure 6B). Each monomer has a mixed α/β structural fold and consists of the cofactor PLP-binding domain (residues 87-336), an N-terminal domain (residues 17-86), and a C-terminal domain (residues 337-441) (Figure 6A). ...
... ∼0.9 Å over 404 Cα atoms) (Shah et al., 1997;Storici et al., 1999), OAT from P. falciparum (r.m.s.d. ∼1.2 Å over 384 Cα atoms) (Vedadi et al., 2007;Jortzik et al., 2010) and OAT from P. yoelii (r.m.s.d. ∼1.0 Å over 369 Cα atoms) (Vedadi et al., 2007). ...
... The cofactor-binding domain of TgOAT non-covalently binds PLP molecule, i.e., PLP does not form the Schiff base with conserve Lys286 (Figures 6A,C). Similarly to homologous OAT structures obtained in complex with the cofactor (Shah et al., 1997;Shen et al., 1998;Storici et al., 1999;Vedadi et al., 2007;Jortzik et al., 2010), PLP interacts with Thr135, Gly136, Ala137, Tyr171, Trp172, Glu224, Asp257, Ile259, Gln260, Lys286, Ser315, and Thr316 ( Figure 6C). Most of these residues are highly conserved among TgOAT homologs and represent a viable, "druggable" pocket (SiteMap score 1.03). ...
Toxoplasma gondii, an Apicomplexan parasite, causes significant morbidity and mortality, including severe disease in immunocompromised hosts and devastating congenital disease, with no effective treatment for the bradyzoite stage. To address this, we used the Tropical Disease Research database, crystallography, molecular modeling, and antisense to identify and characterize a range of potential therapeutic targets for toxoplasmosis. Phosphoglycerate mutase II (PGMII), nucleoside diphosphate kinase (NDK), ribulose phosphate 3-epimerase (RPE), ribose-5-phosphate isomerase (RPI), and ornithine aminotransferase (OAT) were structurally characterized. Crystallography revealed insights into the overall structure, protein oligomeric states and molecular details of active sites important for ligand recognition. Literature and molecular modeling suggested potential inhibitors and druggability. The targets were further studied with vivoPMO to interrupt enzyme synthesis, identifying the targets as potentially important to parasitic replication and, therefore, of therapeutic interest. Targeted vivoPMO resulted in statistically significant perturbation of parasite replication without concomitant host cell toxicity, consistent with a previous CRISPR/Cas9 screen showing PGM, RPE, and RPI contribute to parasite fitness. PGM, RPE, and RPI have the greatest promise for affecting replication in tachyzoites. These targets are shared between other medically important parasites and may have wider therapeutic potential.
... PfOAT (PDB:3LG0, Res. 2.3 Å, (Jortzik et al., 2010)) is active as a homodimer. Based on the comparative sequence alignment with other OATS (Human: HsOAT, PDB: 2OAT, Res. ...
... 1.73 Å (Filippova et al., 2016)) (shown in Fig. 14(A)), we observed high percentage of conserved residues in the active cavity, which is close to the interface of two subunits. Each subunit contains a pyridoxal-phosphate (PLP) binding domain and a substrate binding domain (Jortzik et al., 2010). The PLP binding loop domain (region started from 287 to 293) is strictly conserved in all species. ...
Various reports have shown Cassiarin alkaloids, selective in vitro activities against various strains of Plasmodium falciparum with low cytotoxicity, which indicates their possible candidature as antimalarial drug. However, poor recognition of their protein targets and molecular binding behaviour, certainly limits their exploration as antimalarial drug candidature. To address this, we utilises inverse screening, based on three different docking methodologies in order to find their most putative protein targets. In our study, we screened 1047 protein structures from protein data bank, which belongs to 147 different proteins. Our investigation identified 16 protein targets for Cassiarins. In few cases of identified protein targets, the binding site was poorly studied, which encouraged us to perform comparative sequence and structural studies with their homologous proteins, like as in case of Kelch motif associated protein, Armadillo repeats only protein and Methionine aminopeptidase 1b. In our study, we also found Tryptophanyl-tRNA synthetase and 1-Deoxy-D-Xylose-5-phosphate reductoisomerase proteins are the most common targets for Cassiarins.
