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1.
Int J Mol Sci ; 24(2)2023 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-36674551

RESUMO

The biosynthesis of subunits of rhizobial exopolysaccharides is dependent on glycosyltransferases, which are usually encoded by large gene clusters. PssA is a member of a large family of phosphoglycosyl transferases catalyzing the transfer of a phosphosugar moiety to polyprenol phosphate; thus, it can be considered as priming glycosyltransferase commencing synthesis of the EPS repeating units in Rhizobium leguminosarum. The comprehensive analysis of PssA protein features performed in this work confirmed its specificity for UDP-glucose and provided evidence that PssA is a monotopic inner membrane protein with a reentrant membrane helix rather than a transmembrane segment. The bacterial two-hybrid system screening revealed interactions of PssA with some GTs involved in the EPS octasaccharide synthesis. The distribution of differentially expressed genes in the transcriptome of the ΔpssA mutant into various functional categories indicated complexity of cell response to the deletion, which can mostly be attributed to the lack of exopolysaccharide and downstream effects caused by such deficiency. The block in the EPS biosynthesis at the pssA step, potentially leading to an increased pool of UDP-glucose, is likely to be filtered through to other pathways, and thus the absence of EPS may indirectly affect the expression of proteins involved in these pathways.


Assuntos
Rhizobium leguminosarum , Transferases , Transferases/metabolismo , Rhizobium leguminosarum/metabolismo , Glicosiltransferases/genética , Glicosiltransferases/metabolismo , Fenótipo , Glucose/metabolismo , Difosfato de Uridina/metabolismo , Polissacarídeos Bacterianos/metabolismo , Proteínas de Bactérias/metabolismo
2.
Biochemistry ; 62(1): 109-117, 2023 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-36525630

RESUMO

The Gram-positive pathogen Staphylococcus aureus is a leading cause of antimicrobial resistance related deaths worldwide. Like many pathogens with multidrug-resistant strains, S. aureus contains enzymes that confer resistance through antibiotic modification(s). One such enzyme present in S. aureus is FosB, a Mn2+-dependent l-cysteine or bacillithiol (BSH) transferase that inactivates the antibiotic fosfomycin. fosB gene knockout experiments show that the minimum inhibitory concentration (MIC) of fosfomycin is significantly reduced when the FosB enzyme is not present. This suggests that inhibition of FosB could be an effective method to restore fosfomycin activity. We used high-throughput in silico-based screening to identify small-molecule analogues of fosfomycin that inhibited thiol transferase activity. Phosphonoformate (PPF) was a top hit from our approach. Herein, we have characterized PPF as a competitive inhibitor of FosB from S. aureus (FosBSa) and Bacillus cereus (FosBBc). In addition, we have determined a crystal structure of FosBBc with PPF bound in the active site. Our results will be useful for future structure-based development of FosB inhibitors that can be delivered in combination with fosfomycin in order to increase the efficacy of this antibiotic.


Assuntos
Fosfomicina , Antibacterianos/química , Foscarnet/metabolismo , Foscarnet/farmacologia , Fosfomicina/química , Testes de Sensibilidade Microbiana , Staphylococcus aureus/metabolismo , Transferases/metabolismo , Farmacorresistência Bacteriana , Proteínas de Bactérias/metabolismo
3.
Nat Commun ; 13(1): 7575, 2022 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-36539416

RESUMO

The development of new antibacterial drugs with different mechanisms of action is urgently needed to address antimicrobial resistance. MraY is an essential membrane enzyme required for bacterial cell wall synthesis. Sphaerimicins are naturally occurring macrocyclic nucleoside inhibitors of MraY and are considered a promising target in antibacterial discovery. However, developing sphaerimicins as antibacterials has been challenging due to their complex macrocyclic structures. In this study, we construct their characteristic macrocyclic skeleton via two key reactions. Having then determined the structure of a sphaerimicin analogue bound to MraY, we use a structure-guided approach to design simplified sphaerimicin analogues. These analogues retain potency against MraY and exhibit potent antibacterial activity against Gram-positive bacteria, including clinically isolated drug resistant strains of S. aureus and E. faecium. Our study combines synthetic chemistry, structural biology, and microbiology to provide a platform for the development of MraY inhibitors as antibacterials against drug-resistant bacteria.


Assuntos
Nucleosídeos , Staphylococcus aureus , Nucleosídeos/farmacologia , Nucleosídeos/química , Relação Estrutura-Atividade , Staphylococcus aureus/metabolismo , Antibacterianos/química , Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Transferases/metabolismo
4.
Int J Mol Sci ; 23(20)2022 Oct 19.
Artigo em Inglês | MEDLINE | ID: mdl-36293383

RESUMO

Deodorized garlic (DG) may favor the activity of the antioxidant enzymes and promote the synthesis of hydrogen sulfide (H2S). The objective was to test if DG favors an increase in H2S and if it decreases the oxidative stress caused by lipopolysaccharide (LPS) in rat hearts. A total of 24 rats were divided into 4 groups: Group 1 control (C), Group 2 LPS, Group 3 DG, and Group 4 LPS plus DG. The cardiac mechanical performance (CMP), coronary vascular resistance (CVR), and oxidative stress markers, such as total antioxidant capacity (TAC), glutathione (GSH), selenium (Se), lipid peroxidation (LPO), thiols, hydrogen sulfide (H2S), and the activities and expressions of thioredoxin reductase (TrxR), glutathione peroxidase (GPx), and glutathione-S-transferase (GST), cystathionine synthetase (CBS), cystathionine γ-lyase (CTH), iNOS, and eNOS-p, were analyzed in the heart. Infarct zones in the cardiac tissue were present (p = 0.01). The CMP and CVR decreased and increased (p ≤ 0.05), TAC, GSH, H2S, NO, thiols, and GST activity (p ≤ 0.01) decreased, and LPO and iNOS increased (p ≤ 0.05). The activities and expressions of TrxR, GPx, eNOS-p, CTH, and CBS (p ≤ 0.05) decreased with the LPS treatment; however, DG normalized this effect. DG treatment decreases heart damage caused by LPS through the cross-talk between the H2S and NO systems.


Assuntos
Alho , Sulfeto de Hidrogênio , Selênio , Ratos , Animais , Sulfeto de Hidrogênio/farmacologia , Sulfeto de Hidrogênio/metabolismo , Cistationina gama-Liase/metabolismo , Lipopolissacarídeos/farmacologia , Cistationina beta-Sintase/metabolismo , Alho/metabolismo , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Glutationa Peroxidase/metabolismo , Tiorredoxina Dissulfeto Redutase/metabolismo , Selênio/farmacologia , Estresse Oxidativo , Glutationa/metabolismo , Compostos de Sulfidrila/farmacologia , Transferases/metabolismo , Citidina Monofosfato/metabolismo
5.
ACS Synth Biol ; 11(11): 3699-3705, 2022 Nov 18.
Artigo em Inglês | MEDLINE | ID: mdl-36279362

RESUMO

A key goal of synthetic biology is to enable designed modification of peptides and proteins, both in vivo and in vitro. N- and C-Terminal modification enzymes are crucial in this regard, but there are a few enzymatic options to protect peptide termini. AgeMTPT protects the N-terminus of short peptides with isoprene and the C-terminus as a methyl ester, but its substrate scope is unknown, limiting its application. Here, we investigate the substrate selectivity of the prenyltransferase domain, revealing a requirement for N-terminal aromatic amino acids, but with tolerance for diverse uncharged amino acids in the remaining positions. To demonstrate the potential of the enzyme, substrate selectivity data were used in the enzymatic modification of leu-enkephalin at the critical N-terminal residue. AgeMTPT active site mutagenesis led to an enzyme with expanded substrate scope, including the reverse geranylation of the N-termini of peptides. These data reveal potential applications of enzymatic peptide protection in synthetic biology.


Assuntos
Peptídeos , Transferases , Transferases/metabolismo , Peptídeos/metabolismo , Metiltransferases , Proteínas
6.
Proc Natl Acad Sci U S A ; 119(43): e2202992119, 2022 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-36251991

RESUMO

N-glycosylation is a common posttranslational modification of secreted proteins in eukaryotes. This modification targets asparagine residues within the consensus sequence, N-X-S/T. While this sequence is required for glycosylation, the initial transfer of a high-mannose glycan by oligosaccharyl transferases A or B (OST-A or OST-B) can lead to incomplete occupancy at a given site. Factors that determine the extent of transfer are not well understood, and understanding them may provide insight into the function of these important enzymes. Here, we use mass spectrometry (MS) to simultaneously measure relative occupancies for three N-glycosylation sites on the N-terminal IgV domain of the recombinant glycoprotein, hCEACAM1. We demonstrate that addition is primarily by the OST-B enzyme and propose a kinetic model of OST-B N-glycosylation. Fitting the kinetic model to the MS data yields distinct rates for glycan addition at most sites and suggests a largely stochastic initial order of glycan addition. The model also suggests that glycosylation at one site influences the efficiency of subsequent modifications at the other sites, and glycosylation at the central or N-terminal site leads to dead-end products that seldom lead to full glycosylation of all three sites. Only one path of progressive glycosylation, one initiated by glycosylation at the C-terminal site, can efficiently lead to full occupancy for all three sites. Thus, the hCEACAM1 domain provides an effective model system to study site-specific recognition of glycosylation sequons by OST-B and suggests that the order and efficiency of posttranslational glycosylation is influenced by steric cross-talk between adjoining acceptor sites.


Assuntos
Asparagina , Hexosiltransferases , Asparagina/metabolismo , Glicoproteínas/metabolismo , Glicosilação , Hexosiltransferases/genética , Hexosiltransferases/metabolismo , Manose , Polissacarídeos , Transferases/metabolismo
7.
Int J Mol Sci ; 23(19)2022 Sep 29.
Artigo em Inglês | MEDLINE | ID: mdl-36232831

RESUMO

Plants produce glutathione as a response to the intercellular redox state. Glutathione actively participates in the reactive oxygen species (ROS)-dependent signaling pathway, especially under biotic stress conditions. Most of the glutathione S-transferases (GSTs) are induced in cells during the defense response of plants not only through highly specific glutathione-binding abilities but also by participating in the signaling function. The tau class of GSTs has been reported to be induced as a response under stress conditions. Although several studies have focused on the role of the tau class of GSTs in plant-pathogen interactions, knowledge about their contribution to the response to virus inoculation is still inadequate. Therefore, in this study, the response of Atgstu19 and Atgstu24 knockout mutants to mechanical inoculation of Turnip mosaic virus (TuMV) was examined. The systemic infection of TuMV was more dynamically promoted in Atgstu19 mutants than in wild-type (Col-0) plants, suggesting the role of GSTU19 in TuMV resistance. However, Atgstu24 mutants displayed virus limitation and downregulation of the relative expression of TuMV capsid protein, accompanied rarely by TuMV particles only in vacuoles, and ultrastructural analyses of inoculated leaves revealed the lack of virus cytoplasmic inclusions. These findings indicated that Atgstu24 mutants displayed a resistance-like reaction to TuMV, suggesting that GSTU24 may suppress the plant resistance. In addition, these findings confirmed that GSTU1 and GSTU24 are induced and contribute to the susceptible reaction to TuMV in the Atgstu19-TuMV interaction. However, the upregulation of GSTU19 and GSTU13 highly correlated with virus limitation in the resistance-like reaction in the Atgstu24-TuMV interaction. Furthermore, the highly dynamic upregulation of GST and glutathione reductase (GR) activities resulted in significant induction (between 1 and 14 days post inoculation [dpi]) of the total glutathione pool (GSH + GSSG) in response to TuMV, which was accompanied by the distribution of active glutathione in plant cells. On the contrary, in Atgstu19, which is susceptible to TuMV interaction, upregulation of GST and GR activity only up to 7 dpi symptom development was reported, which resulted in the induction of the total glutathione pool between 1 and 3 dpi. These observations indicated that GSTU19 and GSTU24 are important factors in modulating the response to TuMV in Arabidopsis thaliana. Moreover, it was clear that glutathione is an important component of the regulatory network in resistance and susceptible response of A. thaliana to TuMV. These results help achieve a better understanding of the mechanisms regulating the Arabidopsis-TuMV pathosystem.


Assuntos
Arabidopsis , Potyvirus , Arabidopsis/metabolismo , Proteínas do Capsídeo/metabolismo , Dissulfeto de Glutationa/metabolismo , Glutationa Redutase/metabolismo , Doenças das Plantas/genética , Potyvirus/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Transferases/metabolismo
8.
Int J Mol Sci ; 23(19)2022 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-36233324

RESUMO

Feeding ramie cultivars (Boehmaria nivea L.) are an important feedstock for livestock. Increasing their biomass and improving their nutritional values are essential for animal feeding. Gibberellin (GA3) and ethylene (ETH) are two plant hormones that regulate the growth, development, and metabolism of plants. Herein, we report effects of the GA3 and ETH application on the growth and plant metabolism of feeding ramie in the field. A combination of GA3 and ETH was designed to spray new plants. The two hormones enhanced the growth of plants to produce more biomass. Meanwhile, the two hormones reduced the contents of lignin in leaves and stems, while increased the content of flavonoids in leaves. To understand the potential mechanisms behind these results, we used RNA-seq-based transcriptomics and UPLC-MS/MS-based metabolomics to characterize gene expression and metabolite profiles associated with the treatment of GA3 and ETH. 1562 and 2364 differentially expressed genes (DEGs) were obtained from leaves and stems (treated versus control), respectively. Meanwhile, 99 and 88 differentially accumulated metabolites (DAMs) were annotated from treated versus control leaves and treated versus control stems, respectively. Data mining revealed that both DEGs and DAMs were associated with multiple plant metabolisms, especially plant secondary metabolism. A specific focus on the plant phenylpropanoid pathway identified candidates of DEGs and DEMs that were associated with lignin and flavonoid biosynthesis. Shikimate hydroxycinnamoyl transferase (HCT) is a key enzyme that is involved in the lignin biosynthesis. The gene encoding B. nivea HCT was downregulated in the treated leaves and stems. In addition, genes encoding 4-coumaryl CoA ligase (4CL) and trans-cinnamate 4-monooxygenase (CYP73A), two lignin pathway enzymes, were downregulated in the treated stems. Meanwhile, the reduction in lignin in the treated leaves led to an increase in cinnamic acid and p-coumaryl CoA, two shared substrates of flavonoids that are enhanced in contents. Taken together, these findings indicated that an appropriate combination of GA3 and ETH is an effective strategy to enhance plant growth via altering gene expression and plant secondary metabolism for biomass-enhanced and value-improved feeding ramie.


Assuntos
Boehmeria , Giberelinas , Boehmeria/metabolismo , Cromatografia Líquida , Coenzima A/metabolismo , Etilenos , Flavonoides , Regulação da Expressão Gênica de Plantas , Giberelinas/farmacologia , Hormônios , Ligases/metabolismo , Lignina/metabolismo , Compostos Organofosforados , Reguladores de Crescimento de Plantas/farmacologia , Plantas/metabolismo , Espectrometria de Massas em Tandem , Transcinamato 4-Mono-Oxigenase/genética , Transcinamato 4-Mono-Oxigenase/metabolismo , Transferases/metabolismo
9.
Int J Mol Sci ; 23(19)2022 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-36233329

RESUMO

NAC transcription factors (TFs) could regulate drought stresses in plants; however, the function of NAC TFs in soybeans remains unclear. To unravel NAC TF function, we established that GmNAC12, a NAC TF from soybean (Glycine max), was involved in the manipulation of stress tolerance. The expression of GmNAC12 was significantly upregulated more than 10-fold under drought stress and more than threefold under abscisic acid (ABA) and ethylene (ETH) treatment. In order to determine the function of GmNAC12 under drought stress conditions, we generated GmNAC12 overexpression and knockout lines. The present findings showed that under drought stress, the survival rate of GmNAC12 overexpression lines increased by more than 57% compared with wild-type plants, while the survival rate of GmNAC12 knockout lines decreased by at least 46%. Furthermore, a subcellular localisation analysis showed that the GmNAC12 protein is concentrated in the nucleus of the tobacco cell. In addition, we used a yeast two-hybrid assay to identify 185 proteins that interact with GmNAC12. Gene ontology (GO) and KEGG analysis showed that GmNAC12 interaction proteins are related to chitin, chlorophyll, ubiquitin-protein transferase, and peroxidase activity. Hence, we have inferred that GmNAC12, as a key gene, could positively regulate soybean tolerance to drought stress.


Assuntos
Secas , Soja , Ácido Abscísico/metabolismo , Quitina/metabolismo , Clorofila , Etilenos , Regulação da Expressão Gênica de Plantas , Peroxidases/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Soja/metabolismo , Estresse Fisiológico/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Transferases/metabolismo , Ubiquitinas/metabolismo
10.
J Exp Biol ; 225(19)2022 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-36200468

RESUMO

Migratory birds undergo seasonal changes to muscle biochemistry. Nonetheless, it is unclear to what extent these changes are attributable to the exercise of flight itself versus endogenous changes. Using starlings (Sturnus vulgaris) flying in a wind tunnel, we tested the effects of exercise training, a single bout of flight and dietary lipid composition on pectoralis muscle oxidative enzymes and lipid transporters. Starlings were either unexercised or trained over 2 weeks to fly in a wind tunnel and sampled either immediately following a long flight at the end of this training or after 2 days recovery from this flight. Additionally, they were divided into dietary groups that differed in dietary fatty acid composition (high polyunsaturates versus high monounsaturates) and amount of dietary antioxidant. Trained starlings had elevated (19%) carnitine palmitoyl transferase and elevated (11%) hydroxyacyl-CoA dehydrogenase in pectoralis muscle compared with unexercised controls, but training alone had little effect on lipid transporters. Immediately following a long wind-tunnel flight, starling pectoralis had upregulated lipid transporter mRNA (heart-type fatty acid binding protein, H-FABP, 4.7-fold; fatty acid translocase, 1.9-fold; plasma membrane fatty acid binding protein, 1.6-fold), and upregulated H-FABP protein (68%). Dietary fatty acid composition and the amount of dietary antioxidants had no effect on muscle catabolic enzymes or lipid transporter expression. Our results demonstrate that birds undergo rapid upregulation of catabolic capacity that largely becomes available during flight itself, with minor effects due to training. These effects likely combine with endogenous seasonal changes to create the migratory phenotype observed in the wild.


Assuntos
Estorninhos , Migração Animal/fisiologia , Animais , Antioxidantes/metabolismo , Carnitina/metabolismo , Coenzima A/metabolismo , Proteína 3 Ligante de Ácido Graxo/metabolismo , Proteínas de Ligação a Ácido Graxo/metabolismo , Ácidos Graxos/metabolismo , Músculo Esquelético/metabolismo , Estresse Oxidativo , Oxirredutases/metabolismo , Músculos Peitorais/metabolismo , RNA Mensageiro/genética , Estorninhos/fisiologia , Transferases/metabolismo
11.
Int J Mol Sci ; 23(20)2022 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-36293061

RESUMO

Gamma-glutamyl transferase (GGT) is involved in the progression of atherosclerosis, since its enzymatic activity promotes the generation of reactive oxygen species (ROS). Besides, GGT may act as a prothrombotic factor by inducing tissue factor (TF) expression, independently of its enzymatic activity. The aim of this study was to assess whether GGT-induced TF stimulation was a consequence of binding to toll-like receptor 4 (TLR4) expressed on monocytes, the precursors of macrophages and foam cells which colocalize with GGT activity within atherosclerotic plaques. Experiments were performed in human peripheral blood mononuclear cells (PBMCs), THP-1 cells (a monocytic cellular model), and HEK293 cells, which were genetically modified to study the activation of TLR4. TF procoagulant activity was assessed by a one-stage clotting time test, and TF protein expression was estimated by western blot. Human recombinant (hr) GGT protein increased TF procoagulant activity and protein expression in both PBMCs and THP-1 cells. The GGT-induced TF stimulation was prevented by cellular pretreatment with TLR4/NF-κB inhibitors (LPS-Rs, CLI-095, and BAY-11-7082), and HEK293 cells lacking TLR4 confirmed that TLR4 is essential for GGT-induced activation of NF-κB. In conclusion, hrGGT induced TF expression in monocytes through a cytokine-like mechanism that involved the activation of TLR4/NF-κB signaling.


Assuntos
Tromboplastina , Receptor 4 Toll-Like , Humanos , Receptor 4 Toll-Like/metabolismo , Tromboplastina/metabolismo , Monócitos/metabolismo , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Lipopolissacarídeos/farmacologia , Leucócitos Mononucleares/metabolismo , Células HEK293 , Citocinas/metabolismo , Transferases/metabolismo
12.
Hum Exp Toxicol ; 41: 9603271221129852, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36137816

RESUMO

Lipid metabolism is an important biochemical process in the body. Recent studies have found that environmental endocrine disruptors play an important role in the regulation of lipid metabolism. Bisphenol A (BPA), a common environmental endocrine disruptor, has adverse effects on lipid metabolism, but the mechanism is still unclear. This study aimed to investigate the effects of gestational BPA exposure on hepatic lipid metabolism and its possible mechanism in male offspring. The pregnant Sprague-Dawley rats were exposed to BPA (0, 0.05, 0.5, 5 mg/kg/day) from day 5 to day 19 of gestation to investigate the levels of triglyceride (TG) and total cholesterol (TC), and the expression of liver lipid metabolism-related genes in male offspring rats. The results showed that compared with the control group, the TG and TC levels in serum and liver in BPA-exposed groups was increased. And the expressions of liver fatty acid oxidation related genes, such as peroxisome proliferators-activated receptor α (PPARα) and carnitine palmitoyl transferase 1α (CPT1α), were down-regulated. However, the expressions of fatty acid synthesis related genes, such as sterol regulatory element binding proteins 1 (SREBP-1), acetyl-CoA carboxylase 1 (ACC1), fatty acid synthase (FAS) and stearoyl-CoA desaturase 1 (SCD-1), were up-regulated. The increased protein levels of mTOR and p-CRTC2 suggested that CREB-regulated transcription coactivator 2 (CRTC2) might be an important mediator in the mTOR/SREBP-1 pathway. In conclusion, these results demonstrated that mTOR/CRTC2/SREBP-1 could be affected by gestational BPA exposure, which may involve in the lipid metabolic disorders in later life.


Assuntos
Disruptores Endócrinos , Metabolismo dos Lipídeos , Acetil-CoA Carboxilase/metabolismo , Acetil-CoA Carboxilase/farmacologia , Animais , Compostos Benzidrílicos , Carnitina/farmacologia , Colesterol , Disruptores Endócrinos/toxicidade , Ácido Graxo Sintases/metabolismo , Ácido Graxo Sintases/farmacologia , Ácidos Graxos/farmacologia , Feminino , Fígado , Masculino , PPAR alfa/metabolismo , Proliferadores de Peroxissomos/metabolismo , Proliferadores de Peroxissomos/farmacologia , Fenóis , Gravidez , Ratos , Ratos Sprague-Dawley , Estearoil-CoA Dessaturase/metabolismo , Estearoil-CoA Dessaturase/farmacologia , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/farmacologia , Serina-Treonina Quinases TOR/metabolismo , Transferases/metabolismo , Transferases/farmacologia , Triglicerídeos
13.
Bull Exp Biol Med ; 173(4): 459-463, 2022 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-36058979

RESUMO

Parameters of the lipid metabolism and lipid peroxidation-antioxidant defense system in male adolescents with obesity, representatives of European and Mongoloid ethnicity were evaluated. Discriminant analysis revealed the most informative biochemical parameters for obese male adolescents: glutathione-S-transferase, VLDL cholesterol, glutathione peroxidase, reduced glutathione, and SOD activity for obese male Caucasian adolescents and glutathione-S-transferase, VLDL cholesterol, α-tocopherol, and glutathione peroxidase for Mongoloids obese male adolescents. The use of discriminant analysis allows implementing a differentiated ethnicity-oriented approach to prescribing antioxidant drugs in the complex therapy of obesity.


Assuntos
Antioxidantes , alfa-Tocoferol , Adolescente , Antioxidantes/metabolismo , VLDL-Colesterol , Análise Discriminante , Etnicidade , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Humanos , Peroxidação de Lipídeos , Masculino , Obesidade , Superóxido Dismutase/metabolismo , Transferases/metabolismo
14.
Environ Int ; 169: 107522, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36137426

RESUMO

As new agrochemicals are continuously introduced into agricultural systems, it is essential to investigate their uptake and metabolism by plants to better evaluate their fate and accumulation in crops and the subsequent risks to human exposure. In this study, the uptake and elimination kinetics and transformation of a novel insecticide, cyetpyrafen, in two model crops (lettuce and rice) were first evaluated by hydroponic experiments. Cyetpyrafen was rapidly taken up by plant roots and reached a steady state within 24 h, and it was preferentially accumulated in root parts with root concentration factors up to 2670 mL/g. An uptake mechanism study suggested that root uptake of cyetpyrafen was likely to be dominated by passive diffusion and was difficult to transport via xylem and phloem. Ten phase I and three phase II metabolites of cyetpyrafen were tentatively identified in the hydroponic-plant system through a nontarget screening strategy. The structures of two main metabolites (M-309 and M-391) were confirmed by synthesized standards. The metabolic pathways were proposed including hydroxylation, hydrolysis, dehydrogenation, dehydration and conjugation, which were assumed to be regulated by cytochrome P450, carboxylesterase, glycosyltransferase, glutathione S-transferases and peroxidase. Cyetpyrafen and its main metabolites (M-409, M-309 and M-391) were estimated to be harmful/toxic toward nontarget organisms by theoretical calculation. The high bioaccumulation and extensive transformation of cyetpyrafen highlighted the necessity for systematically assessing the crop uptake and metabolism of new agrochemicals.


Assuntos
Inseticidas , Hidrolases de Éster Carboxílico/metabolismo , Produtos Agrícolas/metabolismo , Glutationa/metabolismo , Glicosiltransferases/metabolismo , Humanos , Inseticidas/metabolismo , Inseticidas/toxicidade , Peroxidases/metabolismo , Raízes de Plantas/metabolismo , Transferases/metabolismo
15.
Commun Biol ; 5(1): 1012, 2022 09 24.
Artigo em Inglês | MEDLINE | ID: mdl-36153380

RESUMO

Anthocyanins, a major class of flavonoids, are important pigments of grape berries. Despite the recent discovery of the genetic cause underlying the loss of color, the metabolomic and molecular responses are unknown. Anthocyanin quantification among diverse berry color muscadines suggests that all genotypes could produce adequate anthocyanin quantities, irrespective of berry color. Transcriptome profiling of contrasting color muscadine genotypes proposes a potential deficiency that occurs within the anthocyanin transport and/or degradation mechanisms and might cause unpigmented berries. Genome-wide association studies highlighted a region on chromosome-4, comprising several genes encoding glutathione S-transferases involved in anthocyanin transport. Sequence comparison among genotypes reveals the presence of two GST4b alleles that differ by substituting the conserved amino acid residue Pro171-to-Leu. Molecular dynamics simulations demonstrate that GST4b2-Leu171 encodes an inactive protein due to modifications within the H-binding site. Population genotyping suggests the recessive inheritance of the unpigmented trait with a GST4b2/2 homozygous. A model defining colorless muscadines' response to the mutation stimulus, avoiding the impact of trapped anthocyanins within the cytoplasm is established.


Assuntos
Antocianinas , Vitis , Aminoácidos/metabolismo , Antocianinas/genética , Flavonoides/análise , Flavonoides/metabolismo , Frutas/genética , Frutas/metabolismo , Estudo de Associação Genômica Ampla , Glutationa/metabolismo , Mutação , Transferases/metabolismo , Vitis/genética , Vitis/metabolismo
16.
Int J Mol Sci ; 23(18)2022 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-36142192

RESUMO

Intercropping with guava (Psidium guajava L.) can assist with the management of Asian citrus psyllid (ACP, Diaphorina citri Kuwayama), the insect vector of the huanglongbing pathogen, in citrus orchards. Sulfur volatiles have a repellent activity and physiological effects, as well as being important components of guava volatiles. In this study, we tested whether the sulfur volatiles emitted by guava plants play a role in plant-plant communications and trigger anti-herbivore activities against ACP in sweet orange plants (Citrus sinensis L. Osbeck). Real-time determination using a proton-transfer-reaction mass spectrometer (PTR-MS) showed that guava plants continuously release methanethiol, dimethyl sulfide (DMS), and dimethyl disulfide (DMDS), and the contents increased rapidly after mechanical damage. The exposure of orange plants to DMDS resulted in the suppression of the developmental performance of ACP. The differential elevation of salicylic acid (SA) levels; the expression of phenylalanine ammonia lyase (PAL), salicylate-O-methyl transferase (SMT), and pathogenesis-related (PR1) genes; the activities of defense-related enzymes PAL, polyphenol oxidase (PPO), and peroxidase (POD); and the total polyphenol content were observed in DMDS-exposed orange plants. The emission of volatiles including myrcene, nonanal, decanal, and methyl salicylate (MeSA) was increased. In addition, phenylpropanoid and flavonoid biosynthesis, and aromatic amino acid (such as phenylalanine, tyrosine, and tryptophan) metabolic pathways were induced. Altogether, our results indicated that DMDS from guava plants can activate defense responses in eavesdropping orange plants and boost their herbivore resistance to ACP, which suggests the possibility of using DMDS as a novel approach for the management of ACP in citrus orchards.


Assuntos
Citrus sinensis , Citrus , Hemípteros , Psidium , Animais , Catecol Oxidase/metabolismo , Citrus/metabolismo , Citrus sinensis/genética , Dissulfetos , Hemípteros/fisiologia , Peroxidases/metabolismo , Fenilalanina/farmacologia , Fenilalanina Amônia-Liase/metabolismo , Doenças das Plantas/genética , Polifenóis/farmacologia , Prótons , Psidium/química , Ácido Salicílico/metabolismo , Enxofre/metabolismo , Transferases/metabolismo , Triptofano/metabolismo , Tirosina/metabolismo
17.
PLoS Negl Trop Dis ; 16(9): e0010766, 2022 09.
Artigo em Inglês | MEDLINE | ID: mdl-36112664

RESUMO

Fasciola hepatica is a trematode parasite that infects animals and humans causing fasciolosis, a worldwide-distributed disease responsible for important economic losses and health problems. This disease is of growing public health concern since parasite isolates resistant to the current treatment (triclabendazole) have increasingly been described. F. hepatica infects its vertebrate host after ingestion of the encysted parasite (metacercariae), which are found in the water or attached to plants. Upon ingestion, newly excysted juveniles of F. hepatica (FhNEJ) emerge in the intestinal lumen and cross the intestinal barrier, reach the peritoneum and migrate to the biliary ducts, where adult worms fully develop. Despite the efforts made to develop new therapeutic and preventive tools, to date, protection against F. hepatica obtained in different animal models is far from optimal. Early events of host-FhNEJ interactions are of paramount importance for the infection progress in fasciolosis, especially those occurring at the host-parasite interface. Nevertheless, studies of FhNEJ responses to the changing host environment encountered during migration across host tissues are still scarce. Here, we set-up an ex vivo model coupled with quantitative SWATH-MS proteomics to study early host-parasite interaction events in fasciolosis. After comparing tegument and somatic fractions from control parasites and FhNEJ that managed to cross a mouse intestinal section ex vivo, a set of parasite proteins whose expression was statistically different were found. These included upregulation of cathepsins L3 and L4, proteolytic inhibitor Fh serpin 2, and a number of molecules linked with nutrient uptake and metabolism, including histone H4, H2A and H2B, low density lipoprotein receptor, tetraspanin, fatty acid binding protein a and glutathione-S-transferase. Downregulated proteins in FhNEJ after gut passage were more numerous than the upregulated ones, and included the heath shock proteins HSP90 and alpha crystallin, amongst others. This study brings new insights into early host-parasite interactions in fasciolosis and sheds light on the proteomic changes in FhNEJ triggered upon excystment and intestinal wall crossing, which could serve to define new targets for the prevention and treatment of this widespread parasitic disease.


Assuntos
Fasciola hepatica , Fasciolíase , alfa-Cristalinas , Animais , Catepsinas , Fasciola hepatica/metabolismo , Fasciolíase/parasitologia , Proteínas de Ligação a Ácido Graxo , Glutationa/metabolismo , Proteínas de Helminto/metabolismo , Histonas/metabolismo , Humanos , Camundongos , Proteômica , Receptores de LDL/metabolismo , Transferases/metabolismo , Triclabendazol , alfa-Cristalinas/metabolismo
18.
J Med Chem ; 65(18): 12386-12402, 2022 09 22.
Artigo em Inglês | MEDLINE | ID: mdl-36069672

RESUMO

An imidazolone → triazolone replacement addressed the limited passive permeability of a series of protein arginine methyl transferase 5 (PRMT5) inhibitors. This increase in passive permeability was unexpected given the increase in the hydrogen bond acceptor (HBA) count and topological polar surface area (TPSA), two descriptors that are typically inversely correlated with permeability. Quantum mechanics (QM) calculations revealed that this unusual effect was due to an electronically driven disconnect between TPSA and 3D-PSA, which manifests in a reduction in overall HBA strength as indicated by the HBA moment descriptor from COSMO-RS (conductor-like screening model for real solvation). HBA moment was subsequently deployed as a design parameter leading to the discovery of inhibitors with not only improved passive permeability but also reduced P-glycoprotein (P-gp) transport. Our case study suggests that hidden polarity as quantified by TPSA-3DPSA can be rationally designed through QM calculations.


Assuntos
Arginina , Antígeno Prostático Específico , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Humanos , Masculino , Permeabilidade , Antígeno Prostático Específico/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Transferases/metabolismo
19.
Int Immunopharmacol ; 112: 109246, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36116153

RESUMO

Vincristine induced peripheral neuropathy (VIPN) is a serious untoward side effect suffered by cancer patients, which still lacks an adequate therapeutic approach. This study examined the alleviating potential of novel methanimine derivatives i.e. (E)-N-(4-nitrobenzylidene)-4-chloro-2-iodobenzamine (KB 9) and (E)-N-(2-methylbenzylidene)-4-chloro-2-iodobenzamine (KB 10) in VIPN. Vincristine was injected in BALB/c mice for 10 days to instigate nociceptive neuropathy. Dynamic and static allodynia, thermal (hot and cold) hyperalgesia were evaluated at 0, 5, 10 and 14 days using cotton brush, Von Frey filament application, hot plate test, acetone drop and cold water respectively. Tumour necrosis factor alpha (TNF-α), interleukin-1ß (IL-1ß), lipid peroxide (LPO), glutathione-S-transferase (GST), catalase (CAT), superoxide dismutase (SOD) and reactive oxygen species (ROS) assays were performed to assess the efficacy of KB9 and KB10 against neuroinflammation and oxidative stress utilizing ELISA, immunohistochemistry and western blot analysis in brain and sciatic nerve tissues. Computational studies were executed to determine the stable binding conformation of both compounds with respect to COX-2 and NF-κB. Interestingly, both compounds substantially reduced protein expression related to neuroinflammation, oxidative stress (LPO, GST, SOD, CAT) and pain (NF-κB, COX-2, IL-1ß and TNF-α). This molecular analysis suggested that the neuroprotective effect of KB9 and KB10 was mediated via regulation of inflammatory signaling pathways. Overall, this study demonstrated that KB9 and KB10 ameliorated vincristine induced neuropathy, through anti-inflammatory, anti-nociceptive and antioxidant mechanisms.


Assuntos
Fármacos Neuroprotetores , Doenças do Sistema Nervoso Periférico , Camundongos , Animais , Vincristina/farmacologia , Catalase/metabolismo , Antioxidantes/uso terapêutico , Interleucina-1beta/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Espécies Reativas de Oxigênio , Fármacos Neuroprotetores/farmacologia , NF-kappa B/metabolismo , Ciclo-Oxigenase 2/metabolismo , Peróxidos Lipídicos/farmacologia , Acetona/farmacologia , Acetona/uso terapêutico , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Doenças do Sistema Nervoso Periférico/patologia , Estresse Oxidativo , Hiperalgesia/tratamento farmacológico , Superóxido Dismutase/metabolismo , Glutationa/metabolismo , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Água , Transferases/metabolismo , Transferases/farmacologia , Transferases/uso terapêutico
20.
Exp Mol Med ; 54(9): 1536-1548, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-36123535

RESUMO

Artesunate (ART) has been indicated as a candidate drug for hepatocellular carcinoma (HCC). Glucosylceramidase (GBA) is required for autophagic degradation. Whether ART regulates autophagic flux by targeting GBA in HCC remains to be defined. Herein, our data demonstrated that the dramatic overexpression of GBA was significantly associated with aggressive progression and short overall survival times in HCC. Subsequent experiments revealed an association between autophagic activity and GBA expression in clinical HCC samples, tumor tissues from a rat model of inflammation-induced HCC and an orthotopic mouse model, and human HCC cell lines. Interestingly, probe labeling identified GBA as an ART target, which was further verified by both a glutathione-S-transferase pulldown assay and surface plasmon resonance analysis. The elevated protein expression of LC3B, the increased numbers of GFP-LC3B puncta and double-membrane vacuoles, and the enhanced expression of SQSTM1/p62 indicated that the degradation of autophagosomes in HCC cells was inhibited by ART treatment. Both the in vitro and in vivo data revealed that autophagosome accumulation through targeting of GBA was responsible for the anti-HCC effects of ART. In summary, this preclinical study identified GBA as one of the direct targets of ART, which may have promising potential to inhibit lysosomal autophagy for HCC therapy.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Animais , Artesunato/farmacologia , Artesunato/uso terapêutico , Autofagia , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Glucosilceramidase/metabolismo , Glucosilceramidase/farmacologia , Glutationa/metabolismo , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Camundongos , Ratos , Proteína Sequestossoma-1 , Transferases/metabolismo , Transferases/farmacologia
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