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1.
Life Sci ; 232: 116604, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31260684

RESUMO

Chronic kidney disease (CKD) patients present L-arginine (L-arg) deficiency and L-arg supplementation has been used as a treatment. In addition, sarcopenia is another common problem in CKD population, resistance training (RT) is one of the conservative strategies developed to prevent CKD progression, and however there are no evidences of a combination of these two strategies to treat CKD outcomes. The aim of this study was to evaluate the effects of oral L-arg supplementation combined with RT in an experimental model of CKD. Twenty-five Munich-Wistar male rats, 8-week-old were divided in 5 groups: Sham (sedentary control), Nx (CKD sedentary), Nx L-arg (CKD sedentary supplemented with 2% of L-arg), Nx RT (CKD exercised) Nx RT + L-arg (CKD exercised and supplemented with 2% of L-arg). CKD model was obtained by a subtotal 5/6 nephrectomy. RT was performed on a ladder climbing, three weekly sessions on non-consecutive days, with an intensity of 70% maximum carrying capacity. They were submitted to RT and/or L-arg supplementation for 10 weeks. There was a significant improvement in muscle strength, renal function, anti-inflammatory cytokines, arginase metabolism and renal fibrosis after RT. However, the combination of RT and L-arg impaired all the improvements promoted by RT alone. The L-arg supplementation alone did not impair renal fibrosis and renal function. In conclusion, RT improved inflammatory balance, muscle strength, renal function and consequently decreased renal fibrosis. Nevertheless, the association with L-arg supplementation prevented all these effects promoted by RT.


Assuntos
Arginina/farmacologia , Condicionamento Físico Animal/fisiologia , Insuficiência Renal Crônica/dietoterapia , Animais , Arginina/metabolismo , Citocinas/metabolismo , Suplementos Nutricionais , Progressão da Doença , Fibrose/metabolismo , Rim/metabolismo , Masculino , Força Muscular/efeitos dos fármacos , Músculo Esquelético/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Condicionamento Físico Animal/métodos , Ratos , Ratos Wistar , Insuficiência Renal Crônica/metabolismo , Treinamento de Resistência/métodos
2.
Arch Virol ; 164(9): 2231-2241, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31177351

RESUMO

Viral persistence alters cellular antiviral activities. Nitric oxide (NO), a highly reactive free radical and a potent antiviral molecule, can inhibit replication of RNA and DNA viruses, but its production and effect during viral persistence are largely unknown. NO synthesis is stimulated in epithelial cells during acute infection with respiratory syncytial virus (RSV) and interferes with viral replication. In this study, we compared the levels of production of NO and expression of its regulatory enzymes, inducible nitric oxide synthase (NOS II) and arginase 1 (Arg-1), during acute and persistent RSV infection in a macrophage cell line to investigate their role in the control and maintenance of viral infection. We observed that NO and NOS II mRNA were induced at higher levels in acutely infected macrophages than in persistently infected macrophages, while the kinetics of NOS II protein expression were similar in both types of infected cultures, except that its disappearance was delayed during acute infection. Thus, NOS II was inducible and expressed at high levels during persistent infection, but production of NO was low relative to acute infection. This was not associated with a lack of enzymatic activity but instead was due to constitutive expression of the Arg-1 enzyme at the mRNA and protein levels, suggesting that arginase restricts availability of L-arginine as a substrate for NOS II to synthesize NO. This hypothesis was supported by showing that arginase enzymatic activity was inhibited in persistently RSV-infected cells by Nω-hydroxy-nor-L-arginine, increasing L-arginine availability in conditioned medium and producing increased levels of nitrites, concurrently with a significant reduction in virus genome replication, implying that Arg-1 overexpression contributes to the maintenance of the RSV genome in the host in persistent infection.


Assuntos
Arginase/metabolismo , Óxido Nítrico/metabolismo , Infecções por Vírus Respiratório Sincicial/virologia , Vírus Sincicial Respiratório Humano/fisiologia , Arginase/genética , Arginina/metabolismo , Regulação para Baixo , Humanos , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Nitritos/metabolismo , Infecções por Vírus Respiratório Sincicial/enzimologia , Infecções por Vírus Respiratório Sincicial/genética , Vírus Sincicial Respiratório Humano/genética , Replicação Viral
3.
Chem Commun (Camb) ; 55(52): 7482-7485, 2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31184653

RESUMO

Protein arginine (Arg) phosphorylation regulates stress responses and virulence in bacteria. With fluorescent activity probes, we show that McsB, a protein Arg kinase, can dephosphorylate phosphoarginine (pArg) residues to produce ATP from ADP, implicating the dynamic control of protein pArg levels by the kinase even without a phosphatase.


Assuntos
Arginina Quinase/metabolismo , Corantes Fluorescentes/química , Difosfato de Adenosina/metabolismo , Trifosfato de Adenosina/metabolismo , Sequência de Aminoácidos , Arginina/análogos & derivados , Arginina/análise , Arginina/química , Arginina/metabolismo , Cromatografia Líquida de Alta Pressão , Compostos Organofosforados/análise , Fosforilação
4.
Cell Mol Life Sci ; 76(15): 2871-2872, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31177294

RESUMO

Protein post-translational modifications (PTMs) have long been a topic of intensive investigation. Covalent additions to the 20 genetically encoded amino acids can alter protein interactions and can even change enzymatic function. In eukarya, PTMs can amplify both the complexity and functional paradigms of the cellular environment. Therefore, PTMs have been of substantial research interest, both for understanding fundamental mechanisms and to provide insight into drug design. Indeed, targeting proteins involved in writing, reading, and erasing PTMs important for human pathologies are some of the most fruitful avenues of drug discovery. In this multi-author review, we explore exciting new work on lysine and arginine methylation, molecular and structural understanding of some of the lysine and arginine methyltransferases (KMTs and PRMTs, respectively), novel insights into nucleic acid methylation, and how the enzymes responsible for writing these PTMs and readers responsible for recognizing these PTMs could be drugged. Here, we introduce the background and the topics covered in this issue.


Assuntos
Processamento de Proteína Pós-Traducional , Arginina/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Histonas/metabolismo , Humanos , Lisina/metabolismo , Metilação , Proteína-Arginina N-Metiltransferases/metabolismo
5.
Phys Chem Chem Phys ; 21(19): 9957-9968, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31041955

RESUMO

The ethylene-forming enzyme (EFE) is a unique member of the Fe(ii)- and 2-oxoglutarate-dependent (Fe/2OG) oxygenases. It converts 2OG into ethylene plus three CO2 molecules (ethylene-forming reaction) and also catalyzes the C5 hydroxylation of l-arginine coupled to the oxidative decarboxylation of 2OG (l-Arg hydroxylation reaction). To uncover the mechanisms of the dual transformations by EFE, quantum mechanical/molecular mechanical (QM/MM) calculations were carried out. Based on the results, a branched mechanism was proposed. An FeII-peroxysuccinate complex with a dissociated CO2 generated through the nucleophilic attack of the superoxo moiety of the Fe-O2 species on the keto carbon of 2OG is the key common intermediate in both reactions. A competition between the subsequent CO2 insertion (a key step in the ethylene-forming pathway) and the O-O bond cleavage (leading to the formation of succinate) governs the product selectivity. The calculated reaction barriers suggested that the CO2 insertion is favored over the O-O bond cleavage. This is consistent with the product preference observed in experiments. By comparison with the results of AsqJ (an Fe/2OG oxygenase that leads to substrate oxidation exclusively), the protein environment was found to be crucial for the selectivity. Further calculations demonstrated that the local electric field of the protein environment in EFE promotes ethylene formation by acting as a charge template, exemplifying the importance of the electrostatic interaction in enzyme catalysis. These findings offer mechanistic insights into the EFE catalysis and provide important clues for better understanding the unique ethylene-forming capability of EFE compared with other Fe/2OG oxygenases.


Assuntos
Arginina/metabolismo , Etilenos/biossíntese , Ácidos Cetoglutáricos/metabolismo , Liases/metabolismo , Arginina/química , Biocatálise , Teoria da Densidade Funcional , Etilenos/química , Hidroxilação , Ácidos Cetoglutáricos/química , Liases/química , Estrutura Molecular , Oxirredução
6.
Anal Chim Acta ; 1068: 111-119, 2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31072472

RESUMO

Arginine/lysine methylation is an important post-translational modification (PTM) involved in DNA repairing, transcriptional regulation, etc. Immunoaffinity enrichment is currently the most widely used methods for the methylproteome analysis. Large-scale analysis of arginine methylation has been realized by using pan-R-methyl antibodies. Unfortunately, pan specific antibodies targeting all three lysine methylation forms are not available. In this study, we presented a novel chromatography-based enrichment method for global methylproteome analysis. The offline multidimensional tandem chromatography combining strong cation exchange (SCX) chromatography, immobilized metal ion affinity chromatography (IMAC) and high-pH reversed-phase chromatography (high-pH RP) was applied in the large-scale analysis of methylproteome. Totally, 860 forms on 765 sites were identified from BEL cells, covering all five arginine/lysine methylation forms. Among them, 27.21% were lysine methylation forms. This technique allows the simultaneous analysis of both arginine and lysine methylation while it has improved performance for the identification of lysine methylation. Therefore, it is a promising strategy for the investigation of biological functions related to methylation.


Assuntos
Cromatografia de Afinidade , Cromatografia por Troca Iônica , Cromatografia de Fase Reversa , Lisina/análise , Proteoma/análise , Arginina/análise , Arginina/metabolismo , Concentração de Íons de Hidrogênio , Lisina/metabolismo , Metilação
7.
Cell Mol Life Sci ; 76(15): 2917-2932, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31123777

RESUMO

Protein arginine methyltransferases (PRMTs) catalyze the methyl transfer to the arginine residues of protein substrates and are classified into three major types based on the final form of the methylated arginine. Recent studies have shown a strong correlation between PRMT expression level and the prognosis of cancer patients. Currently, crystal structures of eight PRMT members have been determined. Kinetic and structural studies have shown that all PRMTs share similar, but unique catalytic and substrate recognition mechanism. In this review, we discuss the structural similarities and differences of different PRMT members, focusing on their overall structure, S-adenosyl-L-methionine-binding pocket, substrate arginine recognition and catalytic mechanisms. Since PRMTs are valuable targets for drug discovery, we also rationally classify the known PRMT inhibitors into five classes and discuss their mechanisms of action at the atomic level.


Assuntos
Proteína-Arginina N-Metiltransferases/metabolismo , Arginina/metabolismo , Sítios de Ligação , Domínio Catalítico , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Humanos , Metilação , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/metabolismo , Proteína-Arginina N-Metiltransferases/antagonistas & inibidores , Especificidade por Substrato
8.
Cell Mol Life Sci ; 76(15): 2933-2956, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31101937

RESUMO

Arginine methylation is a ubiquitous post-translational modification. Three predominant types of arginine-guanidino methylation occur in Eukarya: mono (Rme1/MMA), symmetric (Rme2s/SDMA), and asymmetric (Rme2a/ADMA). Arginine methylation frequently occurs at sites of protein-protein and protein-nucleic acid interactions, providing specificity for binding partners and stabilization of important biological interactions in diverse cellular processes. Each methylarginine isoform-catalyzed by members of the protein arginine methyltransferase family, Type I (PRMT1-4,6,8) and Type II (PRMT5,9)-has unique downstream consequences. Methylarginines are found in ordered domains, domains of low complexity, and in intrinsically disordered regions of proteins-the latter two of which are intimately connected with biological liquid-liquid phase separation. This review highlights discoveries illuminating how arginine methylation affects genome integrity, gene transcription, mRNA splicing and mRNP biology, protein translation and stability, and phase separation. As more proteins and processes are found to be regulated by arginine methylation, its importance for understanding cellular physiology will continue to grow.


Assuntos
Arginina/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Actinas/metabolismo , Cromatina/metabolismo , Reparo do DNA , Histonas/metabolismo , Humanos , Metilação , Processamento de RNA , Ribonucleoproteínas/metabolismo
9.
J Food Sci ; 84(6): 1631-1637, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31059127

RESUMO

The effects of Gynura bicolor aqueous extract (GAE) upon glycemic control, coagulation disorder, lipid accumulation, and glycative, oxidative, and inflammatory stresses in diabetic mice were investigated. Mice were treated with streptozotocin to induce type 1 diabetes. Diabetic mice were divided into four groups, consumed GAE at 0%, 0.25%, 0.5%, or 1%. Normal group consumed standard mouse basal diet. After 8-week treatments, mice were sacrificed after overnight fasting. Results showed that GAE supplement at 0.5% and 1% decreased plasma glucose level and increased plasma insulin level. Diabetes lowered plasma level of protein C and anti-thrombin III; and raised plasminogen activator inhibitor-1 activity and fibrinogen level in plasma. GAE supplement at 0.5% and 1% reversed these alterations. Histological data, assayed by Oil Red O stain, indicated that GAE supplement decreased lipid accumulation in liver. GAE supplement at 0.5% and 1% reduced aldose reductase activity in heart and kidney; and lowered the levels of carboxymethyllysine and pentosidine in plasma and two organs. Diabetes decreased glutathione content, and increased reactive oxygen species, interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α production in heart and kidney. GAE supplement at three test doses reversed these changes. Diabetes upregulated the mRNA expression of p38 and nuclear factor kappa (NF-κ)B in heart and kidney. GAE supplement suppressed the mRNA expression of both p38 and NF-κB. These novel findings suggest that Gynura bicolor is a potent functional food for diabetic prevention or alleviation.


Assuntos
Antidiuréticos/administração & dosagem , Asteraceae/química , Diabetes Mellitus Experimental/tratamento farmacológico , Extratos Vegetais/administração & dosagem , Animais , Arginina/análogos & derivados , Arginina/metabolismo , Glicemia/metabolismo , Diabetes Mellitus Experimental/metabolismo , Glutationa/metabolismo , Humanos , Rim/efeitos dos fármacos , Rim/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Lisina/análogos & derivados , Lisina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/metabolismo , Espécies Reativas de Oxigênio/metabolismo
10.
PLoS Pathog ; 15(4): e1007653, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-31009518

RESUMO

The Gram-negative bacterium Proteus mirabilis is a common cause of catheter-associated urinary tract infections (CAUTI), which can progress to secondary bacteremia. While numerous studies have investigated experimental infection with P. mirabilis in the urinary tract, little is known about pathogenesis in the bloodstream. This study identifies the genes that are important for survival in the bloodstream using a whole-genome transposon insertion-site sequencing (Tn-Seq) approach. A library of 50,000 transposon mutants was utilized to assess the relative contribution of each non-essential gene in the P. mirabilis HI4320 genome to fitness in the livers and spleens of mice at 24 hours following tail vein inoculation compared to growth in RPMI, heat-inactivated (HI) naïve serum, and HI acute phase serum. 138 genes were identified as ex vivo fitness factors in serum, which were primarily involved in amino acid transport and metabolism, and 143 genes were identified as infection-specific in vivo fitness factors for both spleen and liver colonization. Infection-specific fitness factors included genes involved in twin arginine translocation, ammonia incorporation, and polyamine biosynthesis. Mutants in sixteen genes were constructed to validate both the ex vivo and in vivo results of the transposon screen, and 12/16 (75%) exhibited the predicted phenotype. Our studies indicate a role for the twin arginine translocation (tatAC) system in motility, translocation of potential virulence factors, and fitness within the bloodstream. We also demonstrate the interplay between two nitrogen assimilation pathways in the bloodstream, providing evidence that the GS-GOGAT system may be preferentially utilized. Furthermore, we show that a dual-function arginine decarboxylase (speA) is important for fitness within the bloodstream due to its role in putrescine biosynthesis rather than its contribution to maintenance of membrane potential. This study therefore provides insight into pathways needed for fitness within the bloodstream, which may guide strategies to reduce bacteremia-associated mortality.


Assuntos
Amônia/metabolismo , Arginina/metabolismo , Bacteriemia/microbiologia , Poliaminas/metabolismo , Infecções por Proteus/microbiologia , Proteus mirabilis/crescimento & desenvolvimento , Fatores de Virulência/metabolismo , Animais , Bacteriemia/genética , Bacteriemia/metabolismo , Elementos de DNA Transponíveis , Feminino , Aptidão Genética , Sequenciamento de Nucleotídeos em Larga Escala , Camundongos , Camundongos Endogâmicos CBA , Fenótipo , Infecções por Proteus/genética , Infecções por Proteus/metabolismo , Translocação Genética , Fatores de Virulência/genética
11.
Environ Int ; 127: 503-513, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30981021

RESUMO

BACKGROUND: Mechanisms underlying the effects of traffic-related air pollution on people with asthma remain largely unknown, despite the abundance of observational and controlled studies reporting associations between traffic sources and asthma exacerbation and hospitalizations. OBJECTIVES: To identify molecular pathways perturbed following traffic pollution exposures, we analyzed data as part of the Atlanta Commuters Exposure (ACE-2) study, a crossover panel of commuters with and without asthma. METHODS: We measured 27 air pollutants and conducted high-resolution metabolomics profiling on blood samples from 45 commuters before and after each exposure session. We evaluated metabolite and metabolic pathway perturbations using an untargeted metabolome-wide association study framework with pathway analyses and chemical annotation. RESULTS: Most of the measured pollutants were elevated in highway commutes (p < 0.05). From both negative and positive ionization modes, 17,586 and 9087 metabolic features were extracted from plasma, respectively. 494 and 220 unique features were associated with at least 3 of the 27 exposures, respectively (p < 0.05), after controlling confounders and false discovery rates. Pathway analysis indicated alteration of several inflammatory and oxidative stress related metabolic pathways, including leukotriene, vitamin E, cytochrome P450, and tryptophan metabolism. We identified and annotated 45 unique metabolites enriched in these pathways, including arginine, histidine, and methionine. Most of these metabolites were not only associated with multiple pollutants, but also differentially expressed between participants with and without asthma. The analysis indicated that these metabolites collectively participated in an interrelated molecular network centering on arginine metabolism, underlying the impact of traffic-related pollutants on individuals with asthma. CONCLUSIONS: We detected numerous significant metabolic perturbations associated with in-vehicle exposures during commuting and validated metabolites that were closely linked to several inflammatory and redox pathways, elucidating the potential molecular mechanisms of traffic-related air pollution toxicity. These results support future studies of metabolic markers of traffic exposures and the corresponding molecular mechanisms.


Assuntos
Asma/metabolismo , Metaboloma , Poluição Relacionada com o Tráfego , Transportes , Poluição do Ar/análise , Arginina/metabolismo , Asma/induzido quimicamente , Estudos Cross-Over , Hospitalização , Humanos , Metabolômica
12.
Nat Chem Biol ; 15(5): 510-518, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30962626

RESUMO

Protein phosphorylation regulates key processes in all organisms. In Gram-positive bacteria, protein arginine phosphorylation plays a central role in protein quality control by regulating transcription factors and marking aberrant proteins for degradation. Here, we report structural, biochemical, and in vivo data of the responsible kinase, McsB, the founding member of an arginine-specific class of protein kinases. McsB differs in structure and mechanism from protein kinases that act on serine, threonine, and tyrosine residues and instead has a catalytic domain related to that of phosphagen kinases (PhKs), metabolic enzymes that phosphorylate small guanidino compounds. In McsB, the PhK-like phosphotransferase domain is structurally adapted to target protein substrates and is accompanied by a novel phosphoarginine (pArg)-binding domain that allosterically controls protein kinase activity. The identification of distinct pArg reader domains in this study points to a remarkably complex signaling system, thus challenging simplistic views of bacterial protein phosphorylation.


Assuntos
Arginina/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Proteínas Quinases/química , Proteínas Quinases/metabolismo , Arginina/química , Modelos Moleculares , Fosforilação
13.
Ideggyogy Sz ; 72(3-4): 79-88, 2019 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-30957461

RESUMO

Background and purpose: Atrial fibrillation (AF) is the most common arrhythmia diagnosed in clinical practice. We aimed to measure the L-arginine pathway metabolites as well as their ratios in patients with different types of AF or sinus rhythm and to explore the relationship among the markers and clinical variables in the subacute phase of acute ischemic stroke (AIS). Methods: A total of 46 patients with AIS were prospectively enrolled. The patients were divided into three groups based on diagnosis of either sinus rhythm, paroxysmal or permanent AF. Plasma concentration of the L-arginine pathway metabolites were analyzed at post-stroke 24 hours in the three rhythm groups. Besides, clinical variables and laboratory data were recorded. Results: Asymmetric dimetylarginine (ADMA) was significantly higher in patients with permanent AF compared to sinus rhythm (p<0.001). Both ADMA (p<0.001) and symmetric dimethylarginine (SDMA) (p<0.002) at 24 hours were significantly higher among patients with permanent AF compared to those with paroxysmal AF. The L-arginine/SDMA (p<0.031) ratios at 24 hours were significantly higher among patients with sinus rhythm compared to those with permanent AF. ROC analysis also revealed that plasma SDMA cut-off level over 0.639 µmol/L discriminated permanent AF from paroxysmal AF or sinus rhythm with a 90.9% sensitivity and 77.1% specificity. Neutrophil-lymphocyte ratio also showed significantly higher value in individuals with both paroxysmal and permanent AF (p=0.029). Conclusion: Plasma level of SDMA could discriminate permanent from paroxysmal AF in the subacute phase of ischemic stroke. In addition, an increased neutrophil-lymphocyte ratio may suggest inflammatory process in the evolution of atrial fibrillation.


Assuntos
Arginina/sangue , Arginina/metabolismo , Biomarcadores/sangue , Isquemia Encefálica , Acidente Vascular Cerebral/sangue , Fibrilação Atrial , Humanos , Acidente Vascular Cerebral/etiologia
14.
Arch Virol ; 164(6): 1629-1638, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30968211

RESUMO

Trypsin digestion promotes disassembly of GII.3 NoV virus-like particles (VLPs) and binding of VLPs to salivary histo-blood group antigens (HBGAs), but it is not clear which specific regions or residues mediate viral attachment to HBGAs. An earlier study indicated that arginine residues in the predicted surface-exposed loop region are susceptible to trypsin digestion. Here, we introduced single or multiple substitutions of four arginine residues located in the predicted surface-exposed loop region of the GII.3 NoV capsid protein (VP1) and observed their effects on susceptibility to trypsin digestion and binding to HBGAs. All of the mutations in VP1, including single substitutions (R287G, R292G, R296G or R307G) and quadruple substitutions (R287G, R292G, R296G and R307G), permitted successful VLP assembly. After tryptic digestion, all VP1 proteins bearing single point mutations were cleaved, resulting in complete digestion or single fragments with various molecular sizes (27-35 kDa), while the VP1 protein bearing four substitutions was cleaved into two fragments (27-55 kDa). Binding assays using synthetic and salivary HBGAs showed that none of the VP1 mutants (singly or quadruply substituted) exhibited detectable binding to HBGA before or after trypsin cleavage. These results indicated that arginine residues within the predicted surface loop region of GII.3 NoV VP1 were involved directly or indirectly in binding salivary HBGAs and could potentially mediate the HBGA-GII.3 NoV interactions through which host cells become infected.


Assuntos
Antígenos de Grupos Sanguíneos/metabolismo , Proteínas do Capsídeo/química , Proteínas do Capsídeo/metabolismo , Norovirus/metabolismo , Substituição de Aminoácidos , Animais , Arginina/metabolismo , Proteínas do Capsídeo/genética , Humanos , Peso Molecular , Norovirus/química , Norovirus/genética , Ligação Proteica , Saliva/imunologia , Saliva/virologia , Células Sf9 , Tripsina/metabolismo
15.
PLoS Negl Trop Dis ; 13(4): e0007304, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-31017889

RESUMO

The intracellular protozoan parasite Leishmania donovani causes human visceral leishmaniasis. Intracellular L. donovani that proliferate inside macrophage phagolysosomes compete with the host for arginine, creating a situation that endangers parasite survival. Parasites have a sensor that upon arginine deficiency activates an Arginine Deprivation Response (ADR). L. donovani transport arginine via a high-affinity transporter (LdAAP3) that is rapidly up-regulated by ADR in intracellular amastigotes. To date, the sensor and its ligand have not been identified. Here, we show that the conserved amidino group at the distal cap of the arginine side chain is the ligand that activates ADR, in both promastigotes and intracellular amastigotes, and that arginine sensing and transport binding sites are distinct in L. donovani. Finally, upon addition of arginine and analogues to deprived cells, the amidino ligand activates rapid degradation of LdAAP3. This study provides the first identification of an intra-molecular ligand of a sensor that acts during infection.


Assuntos
Sistemas de Transporte de Aminoácidos Básicos/metabolismo , Arginina/metabolismo , Leishmania donovani/metabolismo , Proteínas de Protozoários/metabolismo , Sistemas de Transporte de Aminoácidos Básicos/genética , Arginina/análogos & derivados , Sítios de Ligação , Transporte Biológico , Regulação da Expressão Gênica , Humanos , Leishmania donovani/genética , Macrófagos/parasitologia , Fagossomos/parasitologia , Proteínas de Protozoários/genética , Células THP-1
16.
J Chem Phys ; 150(8): 084106, 2019 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-30823753

RESUMO

Experiments have shown that cholesterol influences the membrane permeability of small molecules, amino acids, and cell-penetrating peptides. However, their exact translocation mechanisms under the influence of cholesterol remain poorly understood. Given the practical importance of cell-penetrating peptides and the existence of varied cholesterol contents in different cell types, it is necessary to examine the permeation of amino acids in cholesterol-containing membranes at atomic level of details. Here, bias-exchange metadynamics simulations were employed to investigate the molecular mechanism of the membrane permeation of two amino acids Arg and Trp important for cell-penetrating peptides in the presence of different concentrations of cholesterol. We found that the free energy barrier of Arg+ (the protonated form) permeation increased linearly as the cholesterol concentration increased, whereas the barrier of Trp permeation had a rapid increase from 0 mol. % to 20 mol. % cholesterol-containing membranes and nearly unchanged from 20 mol. % to 40 mol. % cholesterol-containing membranes. Arg0 becomes slightly more stable than Arg+ at the center of the dipalmitoylphosphatidylcholine (DPPC) membrane with 40 mol. % cholesterol concentrations. As a result, Arg+ has a similar permeability as Trp at 0 mol. % and 20 mol. % cholesterol, but a significantly lower permeability than Trp at 40 mol. % cholesterol. This difference is caused by the gradual reduction of water defects for Arg+ as the cholesterol concentration increases but lack of water defects for Trp in cholesterol-containing membranes. Strong but different orientation dependence between Arg+ and Trp permeations is observed. These results provide an improved microscopic understanding of amino-acid permeation through cholesterol-containing DPPC membrane systems.


Assuntos
Arginina/metabolismo , Membrana Celular/metabolismo , Colesterol/metabolismo , Simulação de Dinâmica Molecular , Triptofano/metabolismo , Membrana Celular/química , Colesterol/química , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Conformação Molecular , Permeabilidade/efeitos dos fármacos , Água/metabolismo
17.
BMC Plant Biol ; 19(1): 108, 2019 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-30894123

RESUMO

BACKGROUND: Nutrition with ammonium (NH4+) can enhance the drought tolerance of rice seedlings in comparison to nutrition with nitrate (NO3-). However, there are still no detailed studies investigating the response of nitric oxide (NO) to the different nitrogen nutrition and water regimes. To study the intrinsic mechanism underpinning this relationship, the time-dependent production of NO and its protective role in the antioxidant defense system of NH4+- or NO3--supplied rice seedlings were studied under water stress. RESULTS: An early NO burst was induced by 3 h of water stress in the roots of seedlings subjected to NH4+ treatment, but this phenomenon was not observed under NO3- treatment. Root oxidative damage induced by water stress was significantly higher for treatment with NO3- than with NH4+ due to reactive oxygen species (ROS) accumulation in the former. Inducing NO production by applying the NO donor 3 h after NO3- treatment alleviated the oxidative damage, while inhibiting the early NO burst by applying the NO scavenger 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide (c-PTIO) increased root oxidative damage in NH4+ treatment. Application of the nitric oxide synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester(L-NAME) completely suppressed NO synthesis in roots 3 h after NH4+ treatment and aggravated water stress-induced oxidative damage. Therefore, the aggravation of oxidative damage by L-NAME might have resulted from changes in the NOS-mediated early NO burst. Water stress also increased the activity of root antioxidant enzymes (catalase, superoxide dismutase, and ascorbate peroxidase). These were further induced by the NO donor but repressed by the NO scavenger and NOS inhibitor in NH4+-treated roots. CONCLUSION: These findings demonstrate that the NOS-mediated early NO burst plays an important role in alleviating oxidative damage induced by water stress by enhancing the antioxidant defenses in roots supplemented with NH4+.


Assuntos
Compostos de Amônio/farmacologia , Desidratação , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico/metabolismo , Oryza/fisiologia , Antioxidantes/metabolismo , Arginina/metabolismo , Citrulina/metabolismo , NG-Nitroarginina Metil Éster/farmacologia , Nitratos/metabolismo , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase/antagonistas & inibidores , Nitroprussiato/farmacologia , Oryza/efeitos dos fármacos , Oxirredução , Proteínas de Plantas/metabolismo , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/metabolismo
18.
Nat Commun ; 10(1): 966, 2019 02 27.
Artigo em Inglês | MEDLINE | ID: mdl-30814497

RESUMO

Current chemical-fuel-driven nanomotors are driven by gas (e.g. H2, O2, NH3) which only provides motion ability, and can produce waste (e.g. Mg(OH)2, Pt). Here, inspired by endogenous biochemical reactions in the human body involving conversion of amino acid L-arginine to nitric oxide (NO) by NO synthase (NOS) or reactive oxygen species (ROS), we report on a nanomotor made of hyperbranched polyamide/L-arginine (HLA). The nanomotor utilizes L-arginine as fuel for the production of NO both as driving force and to provide beneficial effects, including promoting endothelialisation and anticancer effects, along with other beneficial by-products. In addition, the HLA nanomotors are fluorescent and can be used to monitor the movement of nanomotors in vivo in the future. This work presents a zero-waste, self-destroyed and self-imaging nanomotor with potential biological application for the treatment of various diseases in different tissues including blood vessels and tumours.


Assuntos
Nanoestruturas/química , Óxido Nítrico/metabolismo , Arginina/metabolismo , Fontes de Energia Bioelétrica , Bioengenharia , Fluorescência , Células Endoteliais da Veia Umbilical Humana , Humanos , Células MCF-7 , Movimento (Física) , Nanoestruturas/ultraestrutura , Nanotecnologia , Nylons/metabolismo , Espécies Reativas de Oxigênio/metabolismo
19.
Food Funct ; 10(4): 1903-1914, 2019 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-30869672

RESUMO

This study explores the roles of l-arginine (Arg) and N-carbamylglutamate (NCG) supplementation in the diet in intestine damage, energy state, as well as the associated protein kinase signaling pathways activated by AMP in intrauterine growth retarded (IUGR) suckling lambs. A total of 48 newborn Hu lambs with a normal birth weight (CON) and those with IUGR were randomly divided into four groups, CON, IUGR, IUGR + 1% Arg, and IUGR + 0.1% NCG, with 12 animals in each group. All animals were fed for 21 days, from day 7-28, following birth. Our results indicated that the IUGR suckling Hu lambs in the Arg or NCG groups were associated with reduced (P < 0.05) plasma diamine oxidase (DAO) and d-lactic acid levels compared with IUGR suckling lambs. In addition, IUGR suckling Hu lambs in the Arg or NCG group were also linked with a higher (P < 0.05) villous height : crypt depth ratio (VCR), as well as villous height in the duodenum relative to those obtained for IUGR suckling Hu lambs. Relative to IUGR suckling Hu lambs, IUGR suckling Hu lambs in the Arg or NCG groups were found to have higher (P < 0.05) ATP, ADP and TAN contents, and AEC levels, and smaller (P < 0.05) AMP : ATP ratios in the duodenum, jejunum and ileum. Moreover, IUGR suckling Hu lambs in the Arg or NCG group were also linked with higher citrate synthase, isocitrate dehydrogenase and alpha-oxoglutarate dehydrogenase complex activities in the duodenum, jejunum and ileum compared with those found for IUGR suckling Hu lambs (P < 0.05), except for the activity of isocitrate dehydrogenase in the ileum. IUGR suckling Hu lambs in the Arg or NCG group were linked with a lower ratio of pAMPKα/tAMPKα and protein expression of Sirt1 and PGC1α in the ileum relative to those of the IUGR suckling Hu lambs (P < 0.05). Taken together, these findings show that supplementation of NCG and Arg in the diet can ameliorate intestinal injury, improve energy status, motivate key enzyme activities in the tricarboxylic acid (TCA) cycle, and also inhibit the AMP-activated protein kinase signaling pathways in IUGR suckling Hu lambs.


Assuntos
Arginina/metabolismo , Suplementos Nutricionais/análise , Retardo do Crescimento Fetal/metabolismo , Glutamatos/metabolismo , Mucosa Intestinal/metabolismo , Ovinos/metabolismo , Ração Animal/análise , Animais , Arginina/administração & dosagem , Metabolismo Energético/efeitos dos fármacos , Feminino , Retardo do Crescimento Fetal/tratamento farmacológico , Glutamatos/administração & dosagem , Mucosa Intestinal/efeitos dos fármacos , Jejuno/efeitos dos fármacos , Jejuno/metabolismo , Masculino , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Ovinos/crescimento & desenvolvimento , Sirtuína 1/genética , Sirtuína 1/metabolismo
20.
Orphanet J Rare Dis ; 14(1): 63, 2019 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-30832686

RESUMO

BACKGROUND: y+LAT1, encoded by SCL7A7, is the protein mutated in Lysinuric Protein Intolerance (LPI), a rare metabolic disease caused by a defective cationic amino acid (CAA, arginine, lysine, ornithine) transport at the basolateral membrane of intestinal and renal tubular cells. The disease is characterized by protein-rich food intolerance with secondary urea cycle disorder, but symptoms are heterogeneous with lung and immunological complications that are not explainable by the CAA transport defect. With the exception of the Finnish founder mutation (c.895-2A > T, LPIFin), LPI-causative mutations are heterogeneous and genotype-phenotype correlations have not been found. Here we addressed system y+L-mediated arginine uptake in monocytes from three LPI Italian patients and in lymphoblasts carrying the same mutations; in parallel, the genetic defects carried by the patients were reproduced as eGFP-tagged y+LAT1 mutants in transfected CHO cells to define the function and localization protein. RESULTS: System y+L activity is impaired in monocytes isolated from all LPI patients, and in CHO cells transfected with the three eGFP-y+LAT1 mutants, but not in lymphoblasts bearing the same mutations. The analysis of protein localization with confocal microscopy revealed that the eGFP-tagged mutants were retained inside the cytosol, with a pattern of expression quite heterogeneous among the mutants. CONCLUSIONS: The three mutations studied of y+LAT1 transporter result in a defective arginine transport both in ex vivo (monocytes) and in vitro (CHO transfected cells) models, likely caused by the retention of the mutated proteins in the cytosol. The different effect of y+LAT1 mutation on arginine transport in monocytes and lymphoblasts is supposed to be due to the different expression of SLC7A7 mRNA in the two models, supporting the hypothesis that the impact of LPI defect largely depends on the relative abundance of LPI target gene in each cell type.


Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Cadeias Leves da Proteína-1 Reguladora de Fusão/genética , Cadeias Leves da Proteína-1 Reguladora de Fusão/metabolismo , Mutação , Transporte Proteico/genética , Adulto , Animais , Arginina/metabolismo , Células CHO , Células Cultivadas , Criança , Pré-Escolar , Cricetulus , Citosol/metabolismo , Feminino , Humanos , Masculino , Monócitos
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