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Methods in Enzymology 2024Chemoenzymatic synthesis of non-natural terpenes using the promiscuous activity of terpene synthases allows for the expansion of the chemical space of terpenoids with...
Chemoenzymatic synthesis of non-natural terpenes using the promiscuous activity of terpene synthases allows for the expansion of the chemical space of terpenoids with potentially new bioactivities. In this report, we describe protocols for the preparation of a novel aphid attractant, (S)-14,15-dimethylgermacrene D, by exploiting the promiscuity of (S)-germacrene D synthase from Solidago canadensis and using an engineered biocatalytic route to convert prenols to terpenoids. The method uses a combination of five enzymes to carry out the preparation of terpenoid semiochemicals in two steps: (1) diphosphorylation of five or six carbon precursors (prenol, isoprenol and methyl-isoprenol) catalyzed by Plasmodium falciparum choline kinase and Methanocaldococcus jannaschii isopentenyl phosphate kinase to form DMADP, IDP and methyl-IDP, and (2) chain elongation and cyclization catalyzed by Geobacillus stearothermophilus (2E,6E)-farnesyl diphosphate synthase and S. canadensis (S)-germacrene D synthase to produce (S)-germacrene D and (S)-14,15-dimethylgermacrene D. Using this method, new non-natural terpenoids are readily accessible and the approach can be adopted to produce different terpene analogs and terpenoid derivatives with potential novel applications.
Topics: Terpenes; Alkyl and Aryl Transferases; Plasmodium falciparum; Animals; Biocatalysis; Substrate Specificity; Aphids
PubMed: 38942504
DOI: 10.1016/bs.mie.2024.03.015 -
The Science of the Total Environment Jun 2024Arid regions harbor seasonal and permanent wetlands, as biodiversity hotspots crucial for ecosystem services despite harsh conditions. These wetlands, typically...
Arid regions harbor seasonal and permanent wetlands, as biodiversity hotspots crucial for ecosystem services despite harsh conditions. These wetlands, typically dependent on episodic intense rainfall, are understudied compared to their humid counterparts. While the diversity of plants and animals in these wetlands is well-known, the microbial communities remain largely unexplored. To address this knowledge gap, we employed metagenome sequencing technologies to profile protist communities, including pathogenic protozoa, and their associated functional pathways, in sediment of permanent and seasonal arid freshwater wetlands across northern South Africa. Results revealed a core community of protists dominated by phylum Apicomplexa (66.73 %), Euglenazoa (19.03 %), Bacillariophyta (5.44 %), Metamonada (4.65 %), Cryptophyta (1.90 %), and Amoebazoa (1.21 %). Seasonal wetlands showed significantly higher protist diversity compared to permanent wetlands (Shannon index, p = 0.019; Chao1, p = 0.0095). A high abundance and diversity of human and zoonotic pathogenic protists (87.67 %) was observed, with lower levels of photoautotrophs (6.69 %) and limited diversity of phagotrophs (5.64 %). Key photoautotrophs identified included diatoms (Thalassiosiraceae and Phaeodactylaceae) and cryptophytes (genus Hemiselmis and Cryptophyta), with consumers/phagotrophs exhibited a correlation with the bacterial community abundance (r = 0.218, p < 0.001). Pathogenic protozoans identified, include malaria-causing Plasmodium, kinetoplastids (genus Besnoita, Theilleria, Neospora, Toxoplasma, Encephalitozoon, and Babesia) and waterborne protozoans of public health importance (such as Cryptosporidium parvum and Giardia lamblia). Furthermore, the enrichment of pathogenesis-associated pathways (amino acid biosynthesis, peptidoglycan maturation, heme biosynthesis and degradation, and the Calvin-Benson-Bassham cycle), along with virulence gene families identified, highlighted these wetlands as potential reservoirs for infectious diseases. Our results unveil a baseline protist taxonomic and functional composition within arid wetlands, including beneficial and pathogenic protozoa. The close proximity of these wetlands to human activity raises concern for local and transboundary spread of these pathogens. Thus, continued monitoring is vital for disease control and preserving these unique ecosystems.
PubMed: 38942318
DOI: 10.1016/j.scitotenv.2024.174306 -
PLoS Pathogens Jun 2024Plasmodium vivax serological exposure markers (SEMs) have emerged as promising tools for the actionable surveillance and implementation of targeted interventions to...
Plasmodium vivax serological exposure markers (SEMs) have emerged as promising tools for the actionable surveillance and implementation of targeted interventions to accelerate malaria elimination. To determine the dynamic profiles of SEMs in current and past P. vivax infections, we screened and selected 11 P. vivax proteins from 210 putative proteins using protein arrays, with a set of serum samples obtained from patients with acute P. vivax and documented past P. vivax infections. Then we used a murine protein immune model to initially investigate the humoral and memory B cell response involved in the generation of long-lived antibodies. We show that of the 11 proteins, especially C-terminal 42-kDa region of P. vivax merozoite surface protein 1 (PvMSP1-42) induced longer-lasting long-lived antibodies, as these antibodies were detected in individuals infected with P. vivax in the 1960-1970s who were not re-infected until 2012. In addition, we provide a potential mechanism for the maintenance of long-lived antibodies after the induction of PvMSP1-42. The results indicate that PvMSP1-42 induces more CD73+CD80+ memory B cells (MBCs) compared to P. vivax GPI-anchored micronemal antigen (PvGAMA), allowing IgG anti-PvMSP1-42 antibodies to be maintained for a long time.
PubMed: 38941356
DOI: 10.1371/journal.ppat.1012334 -
Metabolomics : Official Journal of the... Jun 2024Burkitt lymphoma (BL) is an aggressive non-Hodgkin lymphoma associated with Plasmodium falciparum and Epstein-Barr virus, both of which affect metabolic pathways. The...
INTRODUCTION
Burkitt lymphoma (BL) is an aggressive non-Hodgkin lymphoma associated with Plasmodium falciparum and Epstein-Barr virus, both of which affect metabolic pathways. The metabolomic patterns of BL is unknown.
MATERIALS AND METHODS
We measured 627 metabolites in pre-chemotherapy treatment plasma samples from 25 male children (6-11 years) with BL and 25 cancer-free area- and age-frequency-matched male controls from the Epidemiology of Burkitt Lymphoma in East African Children and Minors study in Uganda using liquid chromatography-tandem mass spectrometry. Unconditional, age-adjusted logistic regression analysis was used to estimate odds ratios (ORs) and their 95% confidence intervals (CIs) for the BL association with 1-standard deviation increase in the log-metabolite concentration, adjusting for multiple comparisons using false discovery rate (FDR) thresholds and Bonferroni correction.
RESULTS
Compared to controls, levels for 42 metabolite concentrations differed in BL cases (FDR < 0.001), including triacylglyceride (18:0_38:6), alpha-aminobutyric acid (AABA), ceramide (d18:1/20:0), phosphatidylcholine ae C40:6 and phosphatidylcholine C38:6 as the top signals associated with BL (ORs = 6.9 to 14.7, P < 2.4✕10). Two metabolites (triacylglyceride (18:0_38:6) and AABA) selected using stepwise logistic regression discriminated BL cases from controls with an area under the curve of 0.97 (95% CI: 0.94, 1.00).
CONCLUSION
Our findings warrant further examination of plasma metabolites as potential biomarkers for BL risk/diagnosis.
Topics: Humans; Burkitt Lymphoma; Child; Uganda; Male; Case-Control Studies; Metabolomics; Metabolome; Female
PubMed: 38940866
DOI: 10.1007/s11306-024-02130-1 -
Applied and Environmental Microbiology Jun 2024Farnesol salvage, a two-step pathway converting farnesol to farnesyl pyrophosphate (FPP), occurs in bacteria, plants, and animals. This paper investigates the presence...
UNLABELLED
Farnesol salvage, a two-step pathway converting farnesol to farnesyl pyrophosphate (FPP), occurs in bacteria, plants, and animals. This paper investigates the presence of this pathway in fungi. Through bioinformatics, biochemistry, and physiological analyses, we demonstrate its absence in the yeasts and , suggesting a likely absence across fungi. We screened 1,053 fungal genomes, including 34 from , for potential homologs to four genes (, , , and ) known to accomplish farnesol/prenol salvage in other organisms. Additionally, we showed that H-farnesol was not converted to FPP or any other phosphorylated prenol, and exogenous farnesol was not metabolized within 90 minutes at any phase of growth and did not rescue cells from the toxic effects of atorvastatin, but it did elevate the levels of intracellular farnesol (F). All these experiments were conducted with . In sum, we found no evidence for farnesol salvage in fungi.
IMPORTANCE
The absence of farnesol salvage constitutes a major difference in the metabolic capabilities of fungi. In terms of fungal physiology, the lack of farnesol salvage pathways relates to how farnesol acts as a quorum-sensing molecule in and why farnesol should be investigated for use in combination with other known antifungal antibiotics. Its absence is essential for a model (K. W. Nickerson et al., Microbiol Mol Biol Rev 88:e00081-22, 2024), wherein protein farnesylation, protein chaperones, and the unfolded protein response are combined under the unifying umbrella of a cell's intracellular farnesol (F). In terms of human health, farnesol should have at least two different modes of action depending on whether those cells have farnesol salvage. Because animals have farnesol salvage, we can now see the importance of dietary prenols as well as the potential importance of farnesol in treating neurodegenerative diseases such as Parkinson's disease, Alzheimer's disease, and multiple sclerosis.
PubMed: 38940563
DOI: 10.1128/aem.00874-24 -
Scientifica 2024This study provides the design of a microencapsulation formula, physicochemical characterization, and antioxidant, antibacterial, and antiplasmodial activities of...
This study provides the design of a microencapsulation formula, physicochemical characterization, and antioxidant, antibacterial, and antiplasmodial activities of microcapsules. The ethanolic extract of was microencapsulated with chitosan (CHI) and sodium tripolyphosphate (Na-TPP) with various stirring times: 60 minutes (CHI60), 90 minutes (CHI90), and 120 minutes (CHI120). The microcapsules were then observed for physicochemical properties using scanning electron microscopy (SEM) and Fourier-transform infrared spectroscopy (FTIR). The microcapsules were tested for antioxidant activity and antibacterial activity against and using the DPPH (2,2-diphenyl-1-picrylhydrazyl) method. Antiplasmodial bioactivity was assessed through in silico molecular docking. The CHI60 and CHI120 microcapsules exhibited a smaller size and an irregular spherical shape, while the same FTIR profile was observed in CHI90 and CHI120. The bioactivity tests demonstrated that CHI90 exhibited high antibacterial activity against and , while CHI120 exhibited high antioxidant performance. Calcigeroside B and Echinoside B exhibited antiplasmodial activity against the dihydroorotate dehydrogenase (PfDHODH) protein, along with an artemisinin inhibition mechanism. In conclusion, the microcapsules with the CHI90 formula demonstrated the best antibacterial activity, while the CHI120 formula exhibited high antioxidant activity. Two terpenoids, Calcigeroside B and Echinoside B, exhibited the best antiplasmodial activity.
PubMed: 38938546
DOI: 10.1155/2024/5559133 -
Essays in Biochemistry Jun 2024The role of malate dehydrogenase (MDH) in the metabolism of various medically significant protozoan parasites is reviewed. MDH is an NADH-dependent oxidoreductase that...
The role of malate dehydrogenase (MDH) in the metabolism of various medically significant protozoan parasites is reviewed. MDH is an NADH-dependent oxidoreductase that catalyzes interconversion between oxaloacetate and malate, provides metabolic intermediates for both catabolic and anabolic pathways, and can contribute to NAD+/NADH balance in multiple cellular compartments. MDH is present in nearly all organisms; isoforms of MDH from apicomplexans (Plasmodium falciparum, Toxoplasma gondii, Cryptosporidium spp.), trypanosomatids (Trypanosoma brucei, T. cruzi) and anaerobic protozoans (Trichomonas vaginalis, Giardia duodenalis) are presented here. Many parasitic species have complex life cycles and depend on the environment of their hosts for carbon sources and other nutrients. Metabolic plasticity is crucial to parasite transition between host environments; thus, the regulation of metabolic processes is an important area to explore for therapeutic intervention. Common themes in protozoan parasite metabolism include emphasis on glycolytic catabolism, substrate-level phosphorylation, non-traditional uses of common pathways like tricarboxylic acid cycle and adapted or reduced mitochondria-like organelles. We describe the roles of MDH isoforms in these pathways, discuss unusual structural or functional features of these isoforms relevant to activity or drug targeting, and review current studies exploring the therapeutic potential of MDH and related genes. These studies show that MDH activity has important roles in many metabolic pathways, and thus in the metabolic transitions of protozoan parasites needed for success as pathogens.
PubMed: 38938216
DOI: 10.1042/EBC20230075 -
Infection, Genetics and Evolution :... Jun 2024In malaria parasites, the erythrocyte binding-like proteins (EBL) are a family of invasion proteins that are attractive vaccine targets. In the case of Plasmodium vivax,...
In malaria parasites, the erythrocyte binding-like proteins (EBL) are a family of invasion proteins that are attractive vaccine targets. In the case of Plasmodium vivax, the widespread malaria parasite, blood-stage vaccines have been largely focused on a single EBL candidate, the Duffy binding-like domain (DBL) of the Duffy binding protein (DBPII), due to its well-characterized role in the reticulocyte invasion. A novel P. vivax EBL family member, the Erythrocyte binding protein (EBP2, also named EBP or DBP2), binds preferentially to reticulocytes and may mediate an alternative P. vivax invasion pathway. To gain insight into the natural genetic diversity of the DBL domain of EBP2 (region II; EBP2-II), we analyzed ebp2-II gene sequences of 71 P. vivax isolates collected in different endemic settings of the Brazilian Amazon rainforest, where P. vivax is the predominant malaria-associated species. Although most of the substitutions in the ebp2-II gene were non-synonymous and suggested positive selection, the results showed that the DBL domain of the EBP2 was much less polymorphic than that of DBPII. The predominant EBP2 haplotype in the Amazon region corresponded to the C127 reference sequence first described in Cambodia (25% C127-like haplotype). An overview of ebp2-II gene sequences available at GenBank (n = 352) from seven countries (Cambodia, Madagascar, Myanmar, PNG, South Korea, Thailand, Vietnam) confirmed the C127-like haplotype as highly prevalent worldwide. Two out of 43 haplotypes (5 to 20 inferred per country) showed a global frequency of 60%. The results presented here open new avenues of research pursuit while suggesting that a vaccine based on the DBL domain of EBP2 should target a few haplotypes for broad coverage.
PubMed: 38936525
DOI: 10.1016/j.meegid.2024.105628 -
International Journal For Parasitology Jun 2024Establishing an intact intracellular parasitophorous vacuole (PV) that enables efficient nutrient uptake and protein trafficking is essential for the survival and...
Establishing an intact intracellular parasitophorous vacuole (PV) that enables efficient nutrient uptake and protein trafficking is essential for the survival and proliferation of Toxoplasma gondii. Although the PV membrane (PVM)-localized dense granule protein 17 (GRA17) and GRA23 mediate the permeability of the PVM to small molecules, including nutrient uptake and excretion of metabolic by-products, the molecular mechanism by which T. gondii acquires nutrients remains unclear. In this study, we showed that the secreted protein GRA47 contributed to normal PV morphology, PVM permeability to small molecules, growth, and virulence in T. gondii. Co-immunoprecipitation analysis demonstrated potential interaction of GRA47 with GRA72, and the loss of GRA72 affected PV morphology, parasite growth and infectivity. To investigate the biological relationship among GRA47, GRA72, GRA17 and GRA23, attempts were made to construct strains with double gene deletion and overexpressing strains. Only Δgra23Δgra72 was successfully constructed. This strain exhibited a significant increase in the proportion of aberrant PVs compared with the Δgra23 strain. Overexpressing one of the three related GRAs partially rescued PVs with aberrant morphology in Δgra47, Δgra72 and Δgra17, while the expression of the Plasmodium falciparum PVM protein PfExp2, an ortholog of GRA17 and GRA23, fully rescued the PV morphological defect in all three Δgra strains. These results suggest that these three GRA proteins may not be functionally redundant but rather work in different ways to regulate nutrient acquisition. These findings highlight the versatility of the nutrient uptake mechanisms in T. gondii, which may contribute to the parasite's remarkable ability to grow in different cellular niches in a very broad range of hosts.
PubMed: 38936501
DOI: 10.1016/j.ijpara.2024.06.003 -
Journal of Medicinal Food Jun 2024Malaria impedes the ability of primary cells of the immune system to generate an efficacious inflammatory and immune response. Black seed () is a core dietary supplement...
Malaria impedes the ability of primary cells of the immune system to generate an efficacious inflammatory and immune response. Black seed () is a core dietary supplement and food additive in folklore. This study investigated the antioxidant, immunomodulatory, and anti-inflammatory effects of cookies in -infected mice. Aqueous extract of black seed was prepared, and the total phenol and flavonoid contents were determined. The mice were infected with standard inoculum of the strain NK65 . The mice weight and behavioral changes were observed. The mice were fed with the cookies (2.5%, 5%, and 10%) and 10 mg/kg chloroquine for 5 consecutive days after the infection was established. The reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase, catalase, and hematological parameters (red cell indices, leukocytes, and its differentials) in the infected mice were determined. The inflammatory mediators, C-reactive protein (CRP), and myeloperoxidase (MPO) were also assayed. The result revealed that black seed had a total phenol content of 18.73 mgGAE/g and total flavonoid content of 0.36 mgQUE/g. The infected mice treated with cookies showed significantly decreased parasitaemia, MDA, and ROS levels. Furthermore, the results showed significant suppression in proinflammatory mediators (CRP and MPO) levels and enhanced antioxidant status of infected mice treated with . The study suggests that could function as nutraceuticals in the management of infection associated with inflammatory and immunomodulatory disorders.
Topics: Animals; Plasmodium berghei; Malaria; Oxidative Stress; Mice; Nigella sativa; Seeds; Plant Extracts; Male; Antioxidants; Disease Models, Animal; Reactive Oxygen Species; Malondialdehyde; Inflammation; Anti-Inflammatory Agents; Food, Fortified; C-Reactive Protein; Superoxide Dismutase; Humans; Flavonoids; Peroxidase
PubMed: 38935918
DOI: 10.1089/jmf.2023.0181