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1.
Enzyme Microb Technol ; 150: 109881, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34489034

RESUMO

Natural fluorinated products are rare and attract great attention. The de novo fluorometabolites biosynthetic pathway in microbes has been studied. It is revealed that the carbon-fluorine (C-F) bond is formed by an exotic enzyme called fluorinase (FLA) when using fluorine ions and S-adenosyl-l-methionine (SAM) as substrates. However, the resource of the precursor SAM is still elusive. To solve this, a novel methionine adenosyltransferase from Streptomyces xinghaiensis (SxMAT) was identified and characterized. We proved that SAM was enzymatically synthesized by SxMAT, an enzyme that mediated the reaction between adenosine triphosphate (ATP) and l-methionine (l-Met) with 99% diastereoisomeric excess (d.e.) and 80% yield. Such high diastereoselectivity had never been reported before. SxMAT was a Co2+-dependent metalloenzyme. The results showed that the metal cobalt ion contributes to the activity and selectivity of SxMAT. Molecular docking was performed to reveal its catalytic mechanism. The optimal temperature and pH were 55 °C and 8.5, respectively. Lastly, a two-step tandem enzymatic reaction using SxMAT and FLA both from S. xinghaiensis to generate 5'-fluoro-deoxyadenosine (5'-FDA) was performed. This implied that SxMAT may be present in this fluorometabolites biosynthetic route. These results suggested that SxMAT could be a useful biocatalyst for the synthesis of optically pure (S)-S-adenosyl-l-methionine, an important nutraceutical. In addition, SxMAT will probably play an important role in the biosynthetic pathway of fluorinated natural products in bacteria.


Assuntos
Metionina Adenosiltransferase , S-Adenosilmetionina , Vias Biossintéticas , Metionina/metabolismo , Metionina Adenosiltransferase/genética , Metionina Adenosiltransferase/metabolismo , Simulação de Acoplamento Molecular , S-Adenosilmetionina/metabolismo , Streptomyces
2.
Molecules ; 26(17)2021 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-34500777

RESUMO

Human neutrophil elastase (HNE) is a uniquely destructive serine protease with the ability to unleash a wave of proteolytic activity by destroying the inhibitors of other proteases. Although this phenomenon forms an important part of the innate immune response to invading pathogens, it is responsible for the collateral host tissue damage observed in chronic conditions such as chronic obstructive pulmonary disease (COPD), and in more acute disorders such as the lung injuries associated with COVID-19 infection. Previously, a combinatorially selected activity-based probe revealed an unexpected substrate preference for oxidised methionine, which suggests a link to oxidative pathogen clearance by neutrophils. Here we use oxidised model substrates and inhibitors to confirm this observation and to show that neutrophil elastase is specifically selective for the di-oxygenated methionine sulfone rather than the mono-oxygenated methionine sulfoxide. We also posit a critical role for ordered solvent in the mechanism of HNE discrimination between the two oxidised forms methionine residue. Preference for the sulfone form of oxidised methionine is especially significant. While both host and pathogens have the ability to reduce methionine sulfoxide back to methionine, a biological pathway to reduce methionine sulfone is not known. Taken together, these data suggest that the oxidative activity of neutrophils may create rapidly cleaved elastase "super substrates" that directly damage tissue, while initiating a cycle of neutrophil oxidation that increases elastase tissue damage and further neutrophil recruitment.


Assuntos
Imunidade Inata , Elastase de Leucócito/metabolismo , Metionina/análogos & derivados , Neutrófilos/imunologia , Biocatálise , COVID-19/imunologia , COVID-19/patologia , COVID-19/virologia , Domínio Catalítico/genética , Ensaios Enzimáticos , Interações Hospedeiro-Patógeno/imunologia , Humanos , Elastase de Leucócito/antagonistas & inibidores , Elastase de Leucócito/genética , Pulmão/imunologia , Pulmão/patologia , Pulmão/virologia , Metionina/metabolismo , Simulação de Dinâmica Molecular , Infiltração de Neutrófilos , Neutrófilos/enzimologia , Oxirredução/efeitos dos fármacos , Proteólise/efeitos dos fármacos , Doença Pulmonar Obstrutiva Crônica/imunologia , Doença Pulmonar Obstrutiva Crônica/patologia , SARS-CoV-2/imunologia , Especificidade por Substrato/imunologia
3.
Int J Biol Macromol ; 185: 821-831, 2021 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-34216670

RESUMO

Curdlan is a water-insoluble exopolysaccharide produced by Agrobacterium species under nitrogen starvation. The curdlan production in the ΔmdeA, ΔmetA, ΔmetH, and ΔmetZ mutants of methionine biosynthesis pathway of Agrobacterium sp. CGMCC 11546 were significantly impaired. Fermentation profiles of four mutants showed that the consumption of ammonia and sucrose was impaired. Transcriptome analysis of the ΔmetH and ΔmetZ mutants showed that numerous differentially expressed genes involved in the electron transfer chain (ETC) were significantly down-regulated, suggesting that methionine biosynthesis pathway affected the production of energy ATP during the curdlan biosynthesis. Furthermore, metabolomics analysis of the ΔmetH and ΔmetZ mutants showed that ADP and FAD were significantly accumulated, while acetyl-CoA was diminished, suggesting that the impaired curdlan production in the ΔmetH and ΔmetZ mutants might be caused by the insufficient supply of energy ATP. Finally, the addition of both dibasic sodium succinate as a substrate of FAD recycling and methionine significantly restored the curdlan production of four mutants. In conclusion, methionine biosynthesis pathway plays an important role in curdlan biosynthesis in Agrobacterium sp. CGMCC 11546, which affected the sufficient supply of energy ATP from the ETC during the curdlan biosynthesis.


Assuntos
Agrobacterium/fisiologia , Vias Biossintéticas , Metionina/metabolismo , Mutação , beta-Glucanas/metabolismo , Trifosfato de Adenosina/metabolismo , Agrobacterium/genética , Proteínas de Bactérias/genética , Fermentação , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica , Metabolômica , Nitrogênio/metabolismo
4.
Int J Mol Sci ; 22(13)2021 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-34206945

RESUMO

Schizophrenia typically emerges during adolescence, with progression from an ultra-high risk state (UHR) to the first episode of psychosis (FEP) followed by a chronic phase. The detailed pathophysiology of schizophrenia and the factors leading to progression across these stages remain relatively unknown. The current treatment relies on antipsychotics, which are effective for FEP and chronic schizophrenia but ineffective for UHR patients. Antipsychotics modulate dopaminergic and glutamatergic neurotransmission, inflammation, oxidative stress, and membrane lipids pathways. Many of these biological pathways intercommunicate and play a role in schizophrenia pathophysiology. In this context, research of preventive treatment in early stages has explored the antipsychotic effects of omega-3 supplementation in UHR and FEP patients. This review summarizes the action of omega-3 in various biological systems involved in schizophrenia. Similar to antipsychotics, omega-3 supplementation reduces inflammation and oxidative stress, improves myelination, modifies the properties of cell membranes, and influences dopamine and glutamate pathways. Omega-3 supplementation also modulates one-carbon metabolism, the endocannabinoid system, and appears to present neuroprotective properties. Omega-3 has little side effects compared to antipsychotics and may be safely prescribed for UHR patients and as an add-on for FEP patients. This could to lead to more efficacious individualised treatments, thus contributing to precision medicine in psychiatry.


Assuntos
Antipsicóticos/farmacocinética , Ácidos Graxos Ômega-3/metabolismo , Esquizofrenia/metabolismo , Animais , Antipsicóticos/uso terapêutico , Ácido Fólico/metabolismo , Humanos , Metionina/metabolismo , Bainha de Mielina/metabolismo , Esquizofrenia/tratamento farmacológico , Esquizofrenia/prevenção & controle , Transmissão Sináptica
5.
Commun Biol ; 4(1): 725, 2021 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-34117367

RESUMO

Methionine metabolism arises as a key target to elucidate the molecular adaptations underlying animal longevity due to the negative association between longevity and methionine content. The present study follows a comparative approach to analyse plasma methionine metabolic profile using a LC-MS/MS platform from 11 mammalian species with a longevity ranging from 3.5 to 120 years. Our findings demonstrate the existence of a species-specific plasma profile for methionine metabolism associated with longevity characterised by: i) reduced methionine, cystathionine and choline; ii) increased non-polar amino acids; iii) reduced succinate and malate; and iv) increased carnitine. Our results support the existence of plasma longevity features that might respond to an optimised energetic metabolism and intracellular structures found in long-lived species.


Assuntos
Longevidade/fisiologia , Metionina/sangue , Animais , Carnitina/metabolismo , Gatos , Bovinos , Colina/sangue , Colina/metabolismo , Colina/fisiologia , Cistationina/sangue , Cistationina/metabolismo , Cistationina/fisiologia , Cães , Cromatografia Gasosa-Espectrometria de Massas , Cobaias , Cavalos , Humanos , Malatos/sangue , Malatos/metabolismo , Metionina/metabolismo , Metionina/fisiologia , Camundongos , Filogenia , Coelhos , Ratos , Ovinos , Ácido Succínico/sangue , Ácido Succínico/metabolismo , Suínos
6.
Nat Commun ; 12(1): 3486, 2021 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-34108489

RESUMO

The metabolome represents a complex network of biological events that reflects the physiologic state of the organism in health and disease. Additionally, specific metabolites and metabolic signaling pathways have been shown to modulate animal ageing, but whether there are convergent mechanisms uniting these processes remains elusive. Here, we used high resolution mass spectrometry to obtain the metabolomic profiles of canonical longevity pathways in C. elegans to identify metabolites regulating life span. By leveraging the metabolomic profiles across pathways, we found that one carbon metabolism and the folate cycle are pervasively regulated in common. We observed similar changes in long-lived mouse models of reduced insulin/IGF signaling. Genetic manipulation of pathway enzymes and supplementation with one carbon metabolites in C. elegans reveal that regulation of the folate cycle represents a shared causal mechanism of longevity and proteoprotection. Such interventions impact the methionine cycle, and reveal methionine restriction as an underlying mechanism. This comparative approach reveals key metabolic nodes to enhance healthy ageing.


Assuntos
Carbono/metabolismo , Ácido Fólico/metabolismo , Longevidade/fisiologia , Redes e Vias Metabólicas , Animais , Caenorhabditis elegans , Insulina/metabolismo , Longevidade/genética , Redes e Vias Metabólicas/genética , Metaboloma , Metionina/metabolismo , Camundongos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Mutação , Peptídeos/metabolismo , Transdução de Sinais , Tetra-Hidrofolato Desidrogenase/genética , Tetra-Hidrofolato Desidrogenase/metabolismo , Tetra-Hidrofolatos/metabolismo , Timidilato Sintase/genética , Timidilato Sintase/metabolismo
7.
Nutrients ; 13(6)2021 May 24.
Artigo em Inglês | MEDLINE | ID: mdl-34073838

RESUMO

The principal sensing of dietary methionine restriction (MR) occurs in the liver, where it activates multiple transcriptional programs that mediate various biological components of the response. Hepatic Fgf21 is a key target and essential endocrine mediator of the metabolic phenotype produced by dietary MR. The transcription factor, Nfe2l2, is also activated by MR and functions in tandem with hepatic Atf4 to transactivate multiple, antioxidative components of the integrated stress response. However, it is unclear whether the transcriptional responses linked to Nfe2l2 activation by dietary MR are essential to the biological efficacy of the diet. Using mice with liver-specific deletion of Nfe2l2 (Nfe2l2fl/(Alb)) and their floxed littermates (Nfe2l2fl/fl) fed either Control or MR diets, the absence of hepatic Nfe2l2 had no effect on the ability of the MR diet to increase FGF21, reduce body weight and adiposity, and increase energy expenditure. Moreover, the primary elements of the hepatic transcriptome were similarly affected by MR in both genotypes, with the only major differences occurring in induction of the P450-associated drug metabolism pathway and the pentose glucuronate interconversion pathway. The biological significance of these pathways is uncertain but we conclude that hepatic Nfe2l2 is not essential in mediating the metabolic effects of dietary MR.


Assuntos
Fígado/metabolismo , Metionina/deficiência , Fator 2 Relacionado a NF-E2/metabolismo , Adiposidade , Animais , Peso Corporal , Metabolismo Energético , Fatores de Crescimento de Fibroblastos/metabolismo , Genótipo , Masculino , Metionina/administração & dosagem , Metionina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2/genética , Obesidade/dietoterapia , Fenótipo
8.
Viruses ; 13(6)2021 05 21.
Artigo em Inglês | MEDLINE | ID: mdl-34064103

RESUMO

Plant-virus interactions are frequently influenced by elevated temperature, which often increases susceptibility to a virus, a scenario described for potato cultivar Chicago infected with potato virus Y (PVY). In contrast, other potato cultivars such as Gala may have similar resistances to PVY at both normal (22 °C) and high (28 °C) temperatures. To elucidate the mechanisms of temperature-independent antivirus resistance in potato, we analysed responses of Gala plants to PVY at different temperatures using proteomic, transcriptional and metabolic approaches. Here we show that in Gala, PVY infection generally upregulates the accumulation of major enzymes associated with the methionine cycle (MTC) independently of temperature, but that temperature (22 °C or 28 °C) may finely regulate what classes accumulate. The different sets of MTC-related enzymes that are up-regulated at 22 °C or 28 °C likely account for the significantly increased accumulation of S-adenosyl methionine (SAM), a key component of MTC which acts as a universal methyl donor in methylation reactions. In contrast to this, we found that in cultivar Chicago, SAM levels were significantly reduced which correlated with the enhanced susceptibility to PVY at high temperature. Collectively, these data suggest that MTC and its major transmethylation function determines resistance or susceptibility to PVY.


Assuntos
Resistência à Doença , Interações Hospedeiro-Patógeno , Metionina/metabolismo , Doenças das Plantas/virologia , Potyvirus/fisiologia , Solanum tuberosum/metabolismo , Solanum tuberosum/virologia , Cromatografia Líquida , Biologia Computacional/métodos , Temperatura Alta , Redes e Vias Metabólicas , Metilação , Proteínas de Plantas , Espectrometria de Massas em Tandem
9.
Genes (Basel) ; 12(5)2021 05 20.
Artigo em Inglês | MEDLINE | ID: mdl-34065323

RESUMO

Autism spectrum disorder (ASD) is a complex neurodevelopmental condition affecting behavior and communication, presenting with extremely different clinical phenotypes and features. ASD etiology is composite and multifaceted with several causes and risk factors responsible for different individual disease pathophysiological processes and clinical phenotypes. From a genetic and epigenetic side, several candidate genes have been reported as potentially linked to ASD, which can be detected in about 10-25% of patients. Folate gene polymorphisms have been previously associated with other psychiatric and neurodegenerative diseases, mainly focused on gene variants in the DHFR gene (5q14.1; rs70991108, 19bp ins/del), MTHFR gene (1p36.22; rs1801133, C677T and rs1801131, A1298C), and CBS gene (21q22.3; rs876657421, 844ins68). Of note, their roles have been scarcely investigated from a sex/gender viewpoint, though ASD is characterized by a strong sex gap in onset-risk and progression. The aim of the present review is to point out the molecular mechanisms related to intracellular folate recycling affecting in turn remethylation and transsulfuration pathways having potential effects on ASD. Brain epigenome during fetal life necessarily reflects the sex-dependent different imprint of the genome-environment interactions which effects are difficult to decrypt. We here will focus on the DHFR, MTHFR and CBS gene-triad by dissecting their roles in a sex-oriented view, primarily to bring new perspectives in ASD epigenetics.


Assuntos
Transtorno do Espectro Autista/genética , Encéfalo/metabolismo , Epigenoma , Ácido Fólico/metabolismo , Metionina/metabolismo , Animais , Transtorno do Espectro Autista/metabolismo , Feminino , Ácido Fólico/genética , Humanos , Masculino , Metionina/genética , Polimorfismo de Nucleotídeo Único , Fatores Sexuais
10.
Int J Mol Sci ; 22(10)2021 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-34065390

RESUMO

(1) Background: methionine cycle is not only essential for cancer cell proliferation but is also critical for metabolic reprogramming, a cancer hallmark. Hepatic and extrahepatic tissues methionine adenosyltransferases (MATs) are products of two genes, MAT1A and MAT2A that catalyze the formation of S-adenosylmethionine (SAM), the principal biological methyl donor. Glycine N-methyltransferase (GNMT) further utilizes SAM for sarcosine formation, thus it regulates the ratio of SAM:S-adenosylhomocysteine (SAH). (2) Methods: by analyzing the TCGA/GTEx datasets available within GEPIA2, we discovered that breast cancer patients with higher MAT2A had worse survival rate (p = 0.0057). Protein expression pattern of MAT1AA, MAT2A and GNMT were investigated in the tissue microarray in our own cohort (n = 252) by immunohistochemistry. MAT2A C/N expression ratio and cell invasion activity were further investigated in a panel of breast cancer cell lines. (3) Results: GNMT and MAT1A were detected in the cytoplasm, whereas MAT2A showed both cytoplasmic and nuclear immunoreactivity. Neither GNMT nor MAT1A protein expression was associated with patient survival rate in our cohort. Kaplan-Meier survival curves showed that a higher cytoplasmic/nuclear (C/N) MAT2A protein expression ratio correlated with poor overall survival (5 year survival rate: 93.7% vs. 83.3%, C/N ratio ≥ 1.0 vs. C/N ratio < 1.0, log-rank p = 0.004). Accordingly, a MAT2A C/N expression ratio ≥ 1.0 was determined as an independent risk factor by Cox regression analysis (hazard ratio = 2.771, p = 0.018, n = 252). In vitro studies found that breast cancer cell lines with a higher MAT2A C/N ratio were more invasive. (4) Conclusions: the subcellular localization of MAT2A may affect its functions, and elevated MAT2A C/N ratio in breast cancer cells is associated with increased invasiveness. MAT2A C/N expression ratio determined by IHC staining could serve as a novel independent prognostic marker for breast cancer.


Assuntos
Neoplasias da Mama/metabolismo , Metionina Adenosiltransferase/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/patologia , Proliferação de Células/fisiologia , Feminino , Humanos , Linfonodos/metabolismo , Linfonodos/patologia , Metionina/metabolismo , Pessoa de Meia-Idade , Metástase Neoplásica/patologia , Prognóstico
11.
J Biol Chem ; 296: 100797, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34019879

RESUMO

Bacterial methionine biosynthesis can take place by either the trans-sulfurylation route or direct sulfurylation. The enzymes responsible for trans-sulfurylation have been characterized extensively because they occur in model organisms such as Escherichia coli. However, direct sulfurylation is actually the predominant route for methionine biosynthesis across the phylogenetic tree. In this pathway, most bacteria use an O-acetylhomoserine aminocarboxypropyltransferase (MetY) to catalyze the formation of homocysteine from O-acetylhomoserine and bisulfide. Despite the widespread distribution of MetY, this pyridoxal 5'-phosphate-dependent enzyme remains comparatively understudied. To address this knowledge gap, we have characterized the MetY from Thermotoga maritima (TmMetY). At its optimal temperature of 70 °C, TmMetY has a turnover number (apparent kcat = 900 s-1) that is 10- to 700-fold higher than the three other MetY enzymes for which data are available. We also present crystal structures of TmMetY in the internal aldimine form and, fortuitously, with a ß,γ-unsaturated ketimine reaction intermediate. This intermediate is identical to that found in the catalytic cycle of cystathionine γ-synthase (MetB), which is a homologous enzyme from the trans-sulfurylation pathway. By comparing the TmMetY and MetB structures, we have identified Arg270 as a critical determinant of specificity. It helps to wall off the active site of TmMetY, disfavoring the binding of the first MetB substrate, O-succinylhomoserine. It also ensures a strict specificity for bisulfide as the second substrate of MetY by occluding the larger MetB substrate, cysteine. Overall, this work illuminates the subtle structural mechanisms by which homologous pyridoxal 5'-phosphate-dependent enzymes can effect different catalytic, and therefore metabolic, outcomes.


Assuntos
Proteínas de Bactérias/metabolismo , Metionina/metabolismo , Thermotoga maritima/metabolismo , Proteínas de Bactérias/química , Vias Biossintéticas , Cristalografia por Raios X , Cinética , Modelos Moleculares , Thermotoga maritima/química
12.
J Anim Sci ; 99(6)2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-33956968

RESUMO

Two nitrogen balance studies were conducted to evaluate the relative bioavailability values (RBV) of dl-methionine (dl-Met) and dl-methionine hydroxy analog calcium salt (MHA-Ca) to l-methionine (l-Met) as Met sources fed to pigs. In experiment 1, 42 pigs were assigned to 7 treatments feeding with basal diet (BD) formulated to be deficient in Met (0.22% standardized ileal digestible basis) but adequate in other amino acids. Diets included (1) BD, (2) BD + 0.025% dl-Met, (3) BD + 0.050% dl-Met, (4) BD + 0.075% dl-Met, (5) BD + 0.025% l-Met, (6) BD + 0.050% l-Met, and (7) BD + 0.075% l-Met. Increasing levels of l-Met and dl-Met enhanced N retained (g/d) and N retention (% of intake) linearly (P < 0.01). Using a linear slope ratio procedure, a product-to-product RBV of dl-Met compared with l-Met was 94% (95% confidence limits: 65% to 123%) based on N retained expressed as g/d and 99% (95% confidence limits: 70% to 128%) for N retention expressed as % of intake. In experiment 2, 42 pigs were allotted to 7 treatments in another N-balance trial. Diets included (1) BD, (2) BD + 0.025% l-Met, (3) BD + 0.050% l-Met, (4) BD + 0.075% l-Met, (5) BD + 0.030% MHA-Ca, (6) BD + 0.060% MHA-Ca, and (7) BD + 0.089% MHA-Ca. An increase in dietary inclusion rates of l-Met increased (P < 0.01) N retained (g/d) linearly while increasing levels of MHA-Ca had no effects (P > 0.05) on N retained (g/d) and N retention (% of intake). Using linear slope-ratio regression, the RBV of MHA-Ca compared with l-Met was 70% (95% confidence limits: 59% to 81%) on a product-to-product basis or 83% on equimolar basis based on N retained expressed as g/d. Overall, the mean RBV of dl-Met to l-Met of 97% (95% confidence limits cover 100%) indicated that dl-Met and l-Met are equally bioavailable as Met sources in pigs. Compared with l-Met, the RBV of MHA-Ca was lower at 70% (95% confidence limits: 59% to 81%) on a product-to-product basis or 83% on equimolar basis in starter pigs.


Assuntos
Ração Animal , Cálcio , Ração Animal/análise , Animais , Disponibilidade Biológica , Dieta/veterinária , Suplementos Nutricionais , Metionina/análogos & derivados , Metionina/metabolismo , Nitrogênio , Suínos
13.
PLoS One ; 16(4): e0243953, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33930018

RESUMO

Increasing methionine availability in dairy cow diets during the first third of lactation may enhance their performance and health. The aim of this study was to determine the effect of supplementing rumen-protected methionine (Smartamine® M, SM) in a lactation diet with protein and energy levels calculated according to the literature. Seventy-six multiparous Holstein cows (39.1 ± 6.8 kg of milk/d and 65 ± 28 DIM) were assigned to 1 of 2 dietary treatments (38/treatment) according to a randomized complete block design with a 2-wk (covariate) and 10-wk experimental period. Treatments were a basal diet (CON; 3.77 Lys:1Met); and CON + 23 g SM (2.97 Lys:1 Met). Individual milk samples were taken every 2 weeks to determine milk composition. Blood was collected from 24 cows on d+30 d to measure plasma AA levels. Body weight and body condition score (BCS) were measured at the beginning and the end of the experiment. The SM diet promoted higher milk yield (41.7 vs. 40.1 kg/d; P = 0.03). Energy-corrected milk yield (41.0 vs. 38.0 kg/d), milk protein yield (1.30 vs. 1.18 kg/d), milk protein (3.14% vs. 2.97%) and casein (2.39% vs. 2.28%) were also different (P < 0.01) as well as milk fat yield (1.42 vs. 1.29 kg/d; P = 0.02). A trend (P = 0.06) for higher milk fat % (3.41% vs. 3.21%) was observed. Both diets resulted in similar body weight, but CON-fed cows tended (P = 0.08) to have higher BCS. Higher plasma methionine levels were determined with SM compared with CON (29.6 vs. 18.4 µM; P < 0.01), but lysine and histidine were not different. Dietary supplementation of RPM improved productive performance by increasing milk yield and milk components yields, suggesting better dietary AA utilization when Met levels are adjusted in Lys-adequate lactation diets.


Assuntos
Ração Animal , Bovinos/fisiologia , Suplementos Nutricionais , Metionina/metabolismo , Leite/metabolismo , Ração Animal/análise , Criação de Animais Domésticos , Animais , Suplementos Nutricionais/análise , Feminino , Lactação , Metionina/administração & dosagem , Metionina/análise , Leite/química , Proteínas do Leite/análise , Proteínas do Leite/metabolismo , Rúmen/fisiologia
14.
Int J Mol Sci ; 22(6)2021 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-33809777

RESUMO

Since Otto Warburg reported in 1924 that cancer cells address their increased energy requirement through a massive intake of glucose, the cellular energy level has offered a therapeutic anticancer strategy. Methionine restriction (MetR) is one of the most effective approaches for inducing low-energy metabolism (LEM) due to the central position in metabolism of this amino acid. However, no simple in vitro system for the rapid analysis of MetR is currently available, and this study establishes the murine cell line L929 as such a model system. L929 cells react rapidly and efficiently to MetR, and the analysis of more than 150 different metabolites belonging to different classes (amino acids, urea and tricarboxylic acid cycle (TCA) cycles, carbohydrates, etc.) by liquid chromatography/mass spectrometry (LC/MS) defines a metabolic fingerprint and enables the identification of specific metabolites representing normal or MetR conditions. The system facilitates the rapid and efficient testing of potential cancer therapeutic metabolic targets. To date, MS studies of MetR have been performed using organisms and yeast, and the current LC/MS analysis of the intra- and extracellular metabolites in the murine cell line L929 over a period of 5 days thus provides new insights into the effects of MetR at the cellular metabolic level.


Assuntos
Fibroblastos/metabolismo , Metionina/metabolismo , Animais , Morte Celular , Linhagem Celular , Proliferação de Células , Ligantes , Metaboloma , Camundongos , Fator de Necrose Tumoral alfa/metabolismo
15.
Free Radic Biol Med ; 169: 187-215, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33865960

RESUMO

Methionine, either as a free amino acid or included in proteins, can be oxidized into methionine sulfoxide (MetO), which exists as R and S diastereomers. Almost all characterized organisms possess thiol-oxidoreductases named methionine sulfoxide reductase (Msr) enzymes to reduce MetO back to Met. MsrA and MsrB reduce the S and R diastereomers of MetO, respectively, with strict stereospecificity and are found in almost all organisms. Another type of thiol-oxidoreductase, the free-methionine-R-sulfoxide reductase (fRMsr), identified so far in prokaryotes and a few unicellular eukaryotes, reduces the R MetO diastereomer of the free amino acid. Moreover, some bacteria possess molybdenum-containing enzymes that reduce MetO, either in the free or protein-bound forms. All these Msrs play important roles in the protection of organisms against oxidative stress. Fungi are heterotrophic eukaryotes that colonize all niches on Earth and play fundamental functions, in organic matter recycling, as symbionts, or as pathogens of numerous organisms. However, our knowledge on fungal Msrs is still limited. Here, we performed a survey of msr genes in almost 700 genomes across the fungal kingdom. We show that most fungi possess one gene coding for each type of methionine sulfoxide reductase: MsrA, MsrB, and fRMsr. However, several fungi living in anaerobic environments or as obligate intracellular parasites were devoid of msr genes. Sequence inspection and phylogenetic analyses allowed us to identify non-canonical sequences with potentially novel enzymatic properties. Finaly, we identified several ocurences of msr horizontal gene transfer from bacteria to fungi.


Assuntos
Eucariotos , Metionina Sulfóxido Redutases , Eucariotos/metabolismo , Fungos/genética , Metionina/metabolismo , Metionina Sulfóxido Redutases/genética , Metionina Sulfóxido Redutases/metabolismo , Oxirredução , Filogenia
16.
Appl Microbiol Biotechnol ; 105(10): 3919-3929, 2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33929594

RESUMO

Methionine (Met) is an essential amino acid with commercial value in animal feed, human nutrition, and as a chemical precursor. Microbial production of Met has seen intensive investigation towards a more sustainable alternative to the chemical synthesis that currently meets the global Met demand. Indeed, efficient Met biosynthesis has been achieved in genetically modified bacteria that harbor engineered enzymes and streamlined metabolic pathways. Very recently, the export of Met as the final step during its fermentative production has been studied and optimized, primarily through identification and expression of microbial Met efflux transporters. In this mini-review, we summarize the current knowledge on four families of Met export and import transporters that have been harnessed for the production of Met and other valuable biomolecules. These families are discussed with respect to their function, gene regulation, and biotechnological applications. We cover methods for identification and characterization of Met transporters as the basis for the further engineering of these proteins and for exploration of other solute carrier families. The available arsenal of Met transporters from different species and protein families provides blueprints not only for fermentative production but also synthetic biology systems, such as molecular sensors and cell-cell communication systems. KEY POINTS: • Sustainable production of methionine (Met) using microbes is actively explored. • Met transporters of four families increase production yield and specificity. • Further applications include other biosynthetic pathways and synthetic biology.


Assuntos
Biotecnologia , Biologia Sintética , Animais , Fermentação , Humanos , Engenharia Metabólica , Redes e Vias Metabólicas , Metionina/metabolismo
17.
Int J Mol Sci ; 22(6)2021 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-33803786

RESUMO

Cold atmospheric plasma (CAP) has attracted much attention in the fields of biotechnology and medicine owing to its potential utility in clinical applications. Recently accumulating evidence has demonstrated that CAP influences protein structures. However, there remain open questions regarding the molecular mechanisms behind the CAP-induced structural perturbations of biomacromolecules. Here, we investigated the potential effects of CAP irradiation of amyloid ß (Aß), an amyloidogenic protein associated with Alzheimer's disease. Using nuclear magnetic resonance spectroscopy, we observed gradual spectral changes in Aß after a 10 s CAP pretreatment, which also suppressed its fibril formation, as revealed by thioflavin T assay. As per mass spectrometric analyses, these effects were attributed to selective oxidation of the methionine residue (Met) at position 35. Interestingly, this modification occurred when Aß was dissolved into a pre-irradiated buffer, indicating that some reactive species oxidize the Met residue. Our results strongly suggest that the H2O2 generated in the solution by CAP irradiation is responsible for Met oxidation, which inhibits Aß amyloid formation. The findings of the present study provide fundamental insights into plasma biology, giving clues for developing novel applications of CAP.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Gases em Plasma/farmacologia , Sequência de Aminoácidos , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/ultraestrutura , Fluorescência , Espectroscopia de Ressonância Magnética , Metionina/metabolismo , Oxirredução , Agregados Proteicos
18.
Biochem Biophys Res Commun ; 553: 1-8, 2021 05 14.
Artigo em Inglês | MEDLINE | ID: mdl-33752091

RESUMO

BACKGROUND AND AIMS: Hypercholesterolemia is characterized by the elevation of plasma total cholesterol level, especially low-density lipoprotein (LDL) cholesterol. This disease is usually caused by a mutation in genes such as LDL receptor, apolipoprotein B, or proprotein convertase subtilisin/kexin type 9. However, a considerable number of patients with hypercholesterolemia do not have any mutation in these candidate genes. In this study, we examined the difference in the metabolic level between patients with hypercholesterolemia and healthy subjects, and screened the potential biomarkers for this disease. METHODS: Analysis of plasma metabolomics in hypercholesterolemia patients and healthy controls was performed by gas chromatography-mass spectrometry and metabolic correlation networks were constructed using Gephi-0.9.2. RESULTS: First, metabolic profile analysis confirmed the distinct metabolic footprints between the patients and the healthy ones. The potential biomarkers screened by orthogonal partial least-squares discrimination analysis included l-lactic acid, cholesterol, phosphoric acid, d-glucose, urea, and d-allose (Variable importance in the projection > 1). Second, arginine and methionine metabolism were significantly perturbed in hypercholesterolemia patients. Finally, we identified that l-lactic acid, l-lysine, l-glutamine, and l-cysteine had high scores of centrality parameters in the metabolic correlation network. CONCLUSION: Plasma l-lactic acid could be used as a sensitive biomarker for hypercholesterolemia. In addition, arginine biosynthesis and cysteine and methionine metabolism were profoundly altered in patients with hypercholesterolemia.


Assuntos
Biomarcadores/sangue , Biomarcadores/metabolismo , Hipercolesterolemia/sangue , Hipercolesterolemia/metabolismo , Metabolômica , Adolescente , Adulto , Arginina/metabolismo , Estudos de Casos e Controles , Colesterol/metabolismo , Cisteína/metabolismo , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Glucose/metabolismo , Glutamina/metabolismo , Humanos , Ácido Láctico/sangue , Ácido Láctico/metabolismo , Lisina/metabolismo , Masculino , Metionina/metabolismo , Pessoa de Meia-Idade , Ácidos Fosfóricos/metabolismo , Ureia/metabolismo , Adulto Jovem
19.
Mol Genet Metab ; 133(1): 109-112, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33762134

RESUMO

GBA variations are common risk factors for Parkinson's disease (PD), and are found in 21.7% of Ashkenazi PD patients (AJ-PD), 4.23% of them carry an allele, 370Rec, which is different from the common GBA-N370S allele. Using whole-genome-sequencing of 370Rec carriers, N370S carriers, and non-carriers, we characterize the unique 370Rec haplotype in AJ-PDs, and show that it harbors a missense variant replacing the highly conserved methionine-27 with valine in the transmembrane domain of the mitochondrial SLC25A44.


Assuntos
Sistemas de Transporte de Aminoácidos/genética , Predisposição Genética para Doença , Mitocôndrias/genética , Proteínas Mitocondriais/genética , Doença de Parkinson/genética , Proteínas Carreadoras de Solutos/genética , Alelos , Feminino , Genoma Humano/genética , Genótipo , Haplótipos/genética , Heterozigoto , Humanos , Judeus/genética , Masculino , Metionina/metabolismo , Mutação/genética , Doença de Parkinson/patologia , Fatores de Risco , Sequenciamento Completo do Genoma
20.
Biochemistry ; 60(15): 1148-1164, 2021 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-33787242

RESUMO

Proton-coupled electron transfer reactions play critical roles in many aspects of sensory phototransduction. In the case of flavoprotein light sensors, reductive quenching of flavin excited states initiates chemical and conformational changes that ultimately transmit light signals to downstream targets. These reactions generally require neighboring aromatic residues and proton-donating side chains for rapid and coordinated electron and proton transfer to flavin. Although photoreduction of flavoproteins can produce either the anionic (ASQ) or neutral semiquinone (NSQ), the factors that favor one over the other are not well understood. Here we employ a biologically active variant of the light-oxygen-voltage (LOV) domain protein VVD devoid of the adduct-forming Cys residue (VVD-III) to probe the mechanism of flavin photoreduction and protonation. A series of isosteric and conservative residue replacements studied by rate measurements, fluorescence quantum yields, FTIR difference spectroscopy, and molecular dynamics simulations indicate that tyrosine residues facilitate charge recombination reactions that limit sustained flavin reduction, whereas methionine residues facilitate radical propagation and quenching and also gate solvent access for flavin protonation. Replacement of a single surface Met residue with Leu favors formation of the ASQ over the NSQ and desensitizes photoreduction to oxidants. In contrast, increasing site hydrophilicity by Gln substitution promotes rapid NSQ formation and weakens the influence of the redox environment. Overall, the photoreactivity of VVD-III can be understood in terms of redundant electron donors, internal hole quenching, and coupled proton transfer reactions that all depend upon protein conformation, dynamics, and solvent penetration.


Assuntos
Flavinas/metabolismo , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Luz , Metionina/metabolismo , Engenharia de Proteínas , Prótons , Transporte de Elétrons , Proteínas Fúngicas/genética , Simulação de Dinâmica Molecular , Domínios Proteicos
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