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1.
Nature ; 585(7824): 277-282, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32879489

RESUMO

Abnormal epigenetic patterns correlate with effector T cell malfunction in tumours1-4, but the cause of this link is unknown. Here we show that tumour cells disrupt methionine metabolism in CD8+ T cells, thereby lowering intracellular levels of methionine and the methyl donor S-adenosylmethionine (SAM) and resulting in loss of dimethylation at lysine 79 of histone H3 (H3K79me2). Loss of H3K79me2 led to low expression of STAT5 and impaired T cell immunity. Mechanistically, tumour cells avidly consumed methionine and outcompeted T cells for methionine by expressing high levels of the methionine transporter SLC43A2. Genetic and biochemical inhibition of tumour SLC43A2 restored H3K79me2 in T cells, thereby boosting spontaneous and checkpoint-induced tumour immunity. Moreover, methionine supplementation improved the expression of H3K79me2 and STAT5 in T cells, and this was accompanied by increased T cell immunity in tumour-bearing mice and patients with colon cancer. Clinically, tumour SLC43A2 correlated negatively with T cell histone methylation and functional gene signatures. Our results identify a mechanistic connection between methionine metabolism, histone patterns, and T cell immunity in the tumour microenvironment. Thus, cancer methionine consumption is an immune evasion mechanism, and targeting cancer methionine signalling may provide an immunotherapeutic approach.


Assuntos
Sistema L de Transporte de Aminoácidos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Histonas/metabolismo , Metionina/metabolismo , Metilação , Neoplasias/metabolismo , Sistema L de Transporte de Aminoácidos/deficiência , Animais , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Epigênese Genética , Feminino , Histonas/química , Humanos , Camundongos , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/patologia , Receptores de Antígenos de Linfócitos T/metabolismo , Fator de Transcrição STAT5/metabolismo
2.
Nat Commun ; 11(1): 3978, 2020 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-32770044

RESUMO

Methionine restriction, a dietary regimen that protects against metabolic diseases and aging, represses cancer growth and improves cancer therapy. However, the response of different cancer cells to this nutritional manipulation is highly variable, and the molecular determinants of this heterogeneity remain poorly understood. Here we report that hepatocyte nuclear factor 4α (HNF4α) dictates the sensitivity of liver cancer to methionine restriction. We show that hepatic sulfur amino acid (SAA) metabolism is under transcriptional control of HNF4α. Knocking down HNF4α or SAA enzymes in HNF4α-positive epithelial liver cancer lines impairs SAA metabolism, increases resistance to methionine restriction or sorafenib, promotes epithelial-mesenchymal transition, and induces cell migration. Conversely, genetic or metabolic restoration of the transsulfuration pathway in SAA metabolism significantly alleviates the outcomes induced by HNF4α deficiency in liver cancer cells. Our study identifies HNF4α as a regulator of hepatic SAA metabolism that regulates the sensitivity of liver cancer to methionine restriction.


Assuntos
Fator 4 Nuclear de Hepatócito/metabolismo , Neoplasias Hepáticas/metabolismo , Metionina/metabolismo , Animais , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Cisteína/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Fator 4 Nuclear de Hepatócito/genética , Fígado/metabolismo , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Mesoderma/efeitos dos fármacos , Mesoderma/patologia , Redes e Vias Metabólicas/efeitos dos fármacos , Metaboloma/efeitos dos fármacos , Camundongos , Sorafenibe/farmacologia , Transcrição Genética/efeitos dos fármacos
3.
In Vivo ; 34(3 Suppl): 1593-1596, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: covidwho-536995

RESUMO

The Covid-19 pandemic is a world-wide crisis without an effective therapy. While most approaches to therapy are using repurposed drugs that were developed for other diseases, it is thought that targeting the biology of the SARS-CoV-2 virus, which causes Covid-19, can result in an effective therapeutic treatment. The coronavirus RNA cap structure is methylated by two viral methyltransferases that transfer methyl groups from S-adenosylmethionine (SAM). The proper methylation of the virus depends on the level of methionine in the host to form SAM. Herein, we propose to restrict methionine availability by treating the patient with oral recombinant methioninase, aiming to treat Covid-19. By restricting methionine we not only interdict viral replication, which depends on the viral RNA cap methyaltion, but also inhibit the proliferation of the infected cells, which have an increased requirement for methionine. Most importantly, the virally-induced T-cell- and macrophage-mediated cytokine storm, which seems to be a significant cause for Covid-19 deaths, can also be inhibited by restricting methionine, since T-cell and macrophrage activation greatly increases the methionine requirement for these cells. The evidence reviewed here suggests that oral recombinant methioninase could be a promising treatment for coronavirus patients.


Assuntos
Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Liases de Carbono-Enxofre/uso terapêutico , Infecções por Coronavirus/tratamento farmacológico , Metionina/metabolismo , Pneumonia Viral/tratamento farmacológico , Capuzes de RNA/efeitos dos fármacos , Processamento Pós-Transcricional do RNA/efeitos dos fármacos , RNA Viral/efeitos dos fármacos , Administração Oral , Antivirais/administração & dosagem , Proteínas de Bactérias/administração & dosagem , Proteínas de Bactérias/uso terapêutico , Betacoronavirus/fisiologia , Liases de Carbono-Enxofre/administração & dosagem , Ensaios Clínicos como Assunto , Infecções por Coronavirus/complicações , Infecções por Coronavirus/imunologia , Síndrome da Liberação de Citocina/prevenção & controle , Humanos , Ativação Linfocitária/efeitos dos fármacos , Ativação de Macrófagos/efeitos dos fármacos , Metanálise como Assunto , Metilação/efeitos dos fármacos , Pandemias , Pneumonia Viral/complicações , Pneumonia Viral/imunologia , Pseudomonas putida/enzimologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/uso terapêutico , S-Adenosilmetionina/metabolismo , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Replicação Viral/efeitos dos fármacos
4.
In Vivo ; 34(3 Suppl): 1593-1596, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32503816

RESUMO

The Covid-19 pandemic is a world-wide crisis without an effective therapy. While most approaches to therapy are using repurposed drugs that were developed for other diseases, it is thought that targeting the biology of the SARS-CoV-2 virus, which causes Covid-19, can result in an effective therapeutic treatment. The coronavirus RNA cap structure is methylated by two viral methyltransferases that transfer methyl groups from S-adenosylmethionine (SAM). The proper methylation of the virus depends on the level of methionine in the host to form SAM. Herein, we propose to restrict methionine availability by treating the patient with oral recombinant methioninase, aiming to treat Covid-19. By restricting methionine we not only interdict viral replication, which depends on the viral RNA cap methyaltion, but also inhibit the proliferation of the infected cells, which have an increased requirement for methionine. Most importantly, the virally-induced T-cell- and macrophage-mediated cytokine storm, which seems to be a significant cause for Covid-19 deaths, can also be inhibited by restricting methionine, since T-cell and macrophrage activation greatly increases the methionine requirement for these cells. The evidence reviewed here suggests that oral recombinant methioninase could be a promising treatment for coronavirus patients.


Assuntos
Antivirais/uso terapêutico , Betacoronavirus/efeitos dos fármacos , Liases de Carbono-Enxofre/uso terapêutico , Infecções por Coronavirus/tratamento farmacológico , Metionina/metabolismo , Pneumonia Viral/tratamento farmacológico , Capuzes de RNA/efeitos dos fármacos , Processamento Pós-Transcricional do RNA/efeitos dos fármacos , RNA Viral/efeitos dos fármacos , Administração Oral , Antivirais/administração & dosagem , Proteínas de Bactérias/administração & dosagem , Proteínas de Bactérias/uso terapêutico , Betacoronavirus/fisiologia , Liases de Carbono-Enxofre/administração & dosagem , Ensaios Clínicos como Assunto , Infecções por Coronavirus/complicações , Infecções por Coronavirus/imunologia , Síndrome da Liberação de Citocina/prevenção & controle , Humanos , Ativação Linfocitária/efeitos dos fármacos , Ativação de Macrófagos/efeitos dos fármacos , Metanálise como Assunto , Metilação/efeitos dos fármacos , Pandemias , Pneumonia Viral/complicações , Pneumonia Viral/imunologia , Pseudomonas putida/enzimologia , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/uso terapêutico , S-Adenosilmetionina/metabolismo , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Replicação Viral/efeitos dos fármacos
5.
Proc Natl Acad Sci U S A ; 117(23): 13000-13011, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32434918

RESUMO

Extensive studies in prostate cancer and other malignancies have revealed that l-methionine (l-Met) and its metabolites play a critical role in tumorigenesis. Preclinical and clinical studies have demonstrated that systemic restriction of serum l-Met, either via partial dietary restriction or with bacterial l-Met-degrading enzymes exerts potent antitumor effects. However, administration of bacterial l-Met-degrading enzymes has not proven practical for human therapy because of problems with immunogenicity. As the human genome does not encode l-Met-degrading enzymes, we engineered the human cystathionine-γ-lyase (hMGL-4.0) to catalyze the selective degradation of l-Met. At therapeutically relevant dosing, hMGL-4.0 reduces serum l-Met levels to >75% for >72 h and significantly inhibits the growth of multiple prostate cancer allografts/xenografts without weight loss or toxicity. We demonstrate that in vitro, hMGL-4.0 causes tumor cell death, associated with increased reactive oxygen species, S-adenosyl-methionine depletion, global hypomethylation, induction of autophagy, and robust poly(ADP-ribose) polymerase (PARP) cleavage indicative of DNA damage and apoptosis.


Assuntos
Cistationina gama-Liase/farmacologia , Metionina/antagonistas & inibidores , Mutagênese Sítio-Dirigida , Neoplasias da Próstata/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Linhagem Celular Tumoral , Cistationina gama-Liase/genética , Cistationina gama-Liase/isolamento & purificação , Cistationina gama-Liase/uso terapêutico , Dano ao DNA/efeitos dos fármacos , Ensaios Enzimáticos , Humanos , Masculino , Metionina/sangue , Metionina/metabolismo , Camundongos , Poli(ADP-Ribose) Polimerases/metabolismo , Neoplasias da Próstata/sangue , Espécies Reativas de Oxigênio/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/farmacologia , Proteínas Recombinantes/uso terapêutico , Testes de Toxicidade Aguda , Ensaios Antitumorais Modelo de Xenoenxerto
6.
Vet Res ; 51(1): 50, 2020 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-32264939

RESUMO

Two experiments were performed to investigate the effect of different ratios of arginine (Arg) to lysine (Lys) in diets with low (30% Lys; Experiment 1) and high (45% Lys; Experiment 2) methionine (Met) levels on selected metabolic parameters, oxidative and epigenetic DNA damage, and the mechanisms underlying intestinal barrier integrity in turkeys challenged with Clostridium perfringens. In each experiment, 108 one-day-old Hybrid Converter female turkeys were placed in 6 pens (18 birds per pen) and reared for 42 days. At 34, 36 and 37 days of age, half of the birds were subjected to C. perfringens challenge. A 3 × 2 factorial design with three levels of Arg relative to Lys (90, 100 and 110%; Arg90, Arg100 and Arg110, respectively) and C. perfringens infection (-, +) was employed. Challenging birds with C. perfringens increased lipid oxidation and the oxidation and methylation of DNA of intestinal mucosa, and down-regulated the activities of DNA-repairing enzymes. Neither the dietary treatment nor the challenge affected the markers of liver function or metabolism. Arg110 diets with the high Met level induced DNA oxidation and methylation whereas these processes were downregulated in birds fed Arg90 diets. The results indicate that Arg90 diets with high Met levels have a beneficial influence on the indicators of intestinal barrier integrity in turkeys with necrotic enteritis (NE). Despite the analyzed amino acid ratios interacted with the systems responsible for the maintenance of gut integrity in the host organism, this dietary intervention probably enabled birds to cope with NE.


Assuntos
Arginina/metabolismo , Doenças das Aves/fisiopatologia , Infecções por Clostridium/veterinária , Clostridium perfringens/fisiologia , Lisina/metabolismo , Metionina/metabolismo , Ração Animal/análise , Animais , Arginina/administração & dosagem , Proteínas Aviárias/metabolismo , Doenças das Aves/microbiologia , Infecções por Clostridium/microbiologia , Infecções por Clostridium/fisiopatologia , Dano ao DNA , Dieta/veterinária , Suplementos Nutricionais/análise , Epigênese Genética , Expressão Gênica , Lisina/administração & dosagem , Metionina/administração & dosagem , Estresse Oxidativo , Distribuição Aleatória , Proteínas de Junções Íntimas/metabolismo
7.
Obesity (Silver Spring) ; 28(6): 1075-1085, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32348021

RESUMO

OBJECTIVE: Identifying novel approaches to combat obesity is important to improve health span. It was hypothesized that methionine restriction (MR) will induce weight loss in obese mice by reducing adipose tissue mass caused by increased energy expenditure and reprogramming of adipose tissue homeostasis. The roles of adiponectin (ADIPOQ) and fibroblast growth factor 21 (FGF21) during weight loss in MR mice were also tested. METHODS: Diet-induced obese (DIO) male C57BL/6J (wild type), Adipoq-deficient (Adipoq knockout [KO]), Fgf21-KO, and Adipoq-Fgf21 double-KO mice were used. Following a switch to high-fat control (DIO-CF, 60% fat/0.86% methionine) or MR (DIO-MR, 60% fat/0.12% methionine) diet, physiological parameters were measured, and inguinal and perigonadal adipose tissues were examined. RESULTS: Obese mice subjected to MR showed loss of body weight and adiposity, increased energy expenditure, and improved glucose tolerance that were independent of the actions of ADIPOQ and FGF21. MR induced reduction of circulating lipids, glucose, insulin, leptin, and insulin like growth factor 1 and increased ß-hydroxybutyrate, ADIPOQ, and FGF21 concentrations. In fat, MR upregulated protein levels of adipose triglyceride lipase, apoptosis-inducing factor, lysosomal-associated membrane proteins 1 and 2, autophagy-related protein 5, beclin-1, and light chain 3B I and II. CONCLUSIONS: MR reduction of adipose tissue mass in obese mice is associated with elevated lipolysis, apoptosis, and autophagy and occurs independently of the actions of ADIPOQ and FGF21.


Assuntos
Adiponectina/metabolismo , Adiposidade/fisiologia , Fatores de Crescimento de Fibroblastos/metabolismo , Metionina/metabolismo , Camundongos Obesos/genética , Perda de Peso/fisiologia , Animais , Masculino , Camundongos
8.
J Dairy Sci ; 103(6): 5668-5683, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32278559

RESUMO

The present review focuses on methyl donor metabolism and nutrition in the periparturient and lactating dairy cow. Methyl donors are involved in one-carbon metabolism, which includes the folate and Met cycles. These cycles work in unison to support lipid, nucleotide, and protein synthesis, as well as methylation reactions and the maintenance of redox status. A key feature of one-carbon metabolism is the multi-step conversion of tetrahydrofolate to 5-methyltetrahyrofolate. Homocysteine and 5-methyltetrahyrofolate are utilized by vitamin B12-dependent Met synthase to couple the folate and Met cycles and generate Met. Methionine may also be remethylated from choline-derived betaine under the action of betaine hydroxymethyltransferase. Regardless, Met is converted within the Met cycle to S-adenosylmethionine, which is universally utilized in methyl-group transfer reactions including the synthesis of phosphatidylcholine. Homocysteine may also enter the transsulfuration pathway to generate glutathione or taurine for scavenging of reactive oxygen metabolites. In the transition cow, a high demand exists for compounds with a labile methyl group. Limited methyl group supply may contribute to inadequate hepatic phosphatidylcholine synthesis and hepatic triglyceride export, systemic oxidative stress, and compromised milk production. To minimize the perils associated with methyl donor deficiency, the peripartum cow relies on de novo methylneogenesis from tetrahydrofolate. In addition, dietary supplementation of rumen-protected folic acid, vitamin B12, Met, choline, and betaine are potential nutritional approaches to target one-carbon pools and improve methyl donor balance in transition cows. Such strategies have merit considering research demonstrating their ability to improve milk production efficiency, milk protein synthesis, hepatic health, and immune response. This review aims to summarize the current understanding of folic acid, vitamin B12, Met, choline, and betaine utilization in the dairy cow. Methyl donor co-supplementation, fatty acid feeding strategies that may optimize methyl donor supplementation efficacy, and potential epigenetic mechanisms are also considered.


Assuntos
Fenômenos Fisiológicos da Nutrição Animal , Bovinos/fisiologia , Dieta/veterinária , Tetra-Hidrofolatos/metabolismo , Animais , Feminino , Ácido Fólico/metabolismo , Metionina/metabolismo
9.
Ecotoxicol Environ Saf ; 194: 110338, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32135376

RESUMO

2,2',3,5',6-Pentachlorobiphenyl (PCB95) is known as a persistent pollutant that was found in eggs in China. PCB 95 can be metabolized into OH-PCB95 and MeO-PCB95 in liver microsomes. However, the toxicity and its mechanism of PCB95 or its metabolites have been little studied on laying hens. Herein, chicken embryo liver cells of laying hens were selected and treated with different levels of PCB95 and its two metabolites, and the EC50 of PCB95, OH-PCB95, MeO-PCB95 was 80.85, 4.81 and 107.04 µg/mL respectively, indicating that OH-PCB95 is much more cytotoxic than PCB95 or MeO-PCB95. Targeted metabolomics was further used to study the effects of the parent compound and its metabolites on cell metabolism. The results showed that four primary types of glycerophospholipids were down-regulated after exposure to PCB95 and its metabolites, especially PE and PS (60% more than the control for PCB95, 40% for OH-PCB95, and less than 40% for MeO-PCB95). KEGG pathway analysis based on amino acid metabolism showed that PCB95 may mainly interfere with the amino acids involved in immune regulation (phenylalanine and tyrosine), and OH-PCB95 may be associated with genetic disoders (cysteine, methionine and purine metabolism). However, the metabolic pathways induced by MeO-PCB95 are quite different from those induced by PCB95 and OH-PCB95, affecting mainly D-glutamine and D-glutamate metabolism, alanine and glutamate metabolism, and arginine and proline metabolism; these pathways mainly regulate the elimination of excess purines and are involved in the synthesis of the amino acids required by cells. These results showed that OH-PCB95 has the highest toxicity on chicken embryo liver cells and MeO-PCB95 could be a detoxification product of PCB95 and OH-PCB95. This study contributes to the understanding of the different effects of PCB95 and its metabolites on cellular metabolism, and the data are helpful in evaluating the hepatotoxic effects of these compounds.


Assuntos
Poluentes Ambientais/toxicidade , Bifenilos Policlorados/toxicidade , Aminoácidos/metabolismo , Animais , Embrião de Galinha , Galinhas/metabolismo , China , Ovos , Poluentes Ambientais/metabolismo , Feminino , Hepatócitos/metabolismo , Fígado/metabolismo , Redes e Vias Metabólicas , Metabolômica , Metionina/metabolismo , Microssomos Hepáticos/metabolismo , Testes de Toxicidade
10.
J Biosci Bioeng ; 130(1): 14-19, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32217026

RESUMO

Here, proteins involved in sulfur-containing amino acid uptake in Escherichia coli strains were investigated with the aim of applying the findings in fermentative amino acid production. A search of genes in an l-methionine auxotrophic strain library suggested YecSC as the putative transporter of l-cystathionine. l-Methionine production increased by 15% after amplification of yecSC in producer strains. A candidate protein responsible for l-cysteine uptake was also found by experimentation with multicopy suppressor E. coli strains that recovered from growth defects caused by l-cysteine auxotrophy. Based on the results of an uptake assay, growth using l-cysteine as a sole sulfur source, and sensitivity to l-cysteine toxicity, we proposed that YeaN is an l-cysteine transporter. l-Cysteine production increased by 50% as a result of disrupting yeaN in producer strain. The study of amino acid transporters is valuable to industrialized amino acid production and also sheds light on the role of these transporters in sulfur assimilation.


Assuntos
Cistationina/metabolismo , Cisteína/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Metionina/metabolismo , Enxofre/metabolismo , Transporte Biológico , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Fermentação , Proteínas de Membrana Transportadoras/metabolismo , Engenharia Metabólica
11.
Proc Natl Acad Sci U S A ; 117(14): 7755-7763, 2020 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-32193337

RESUMO

Methionine metabolism is critical for the maintenance of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) pluripotency. However, little is known about the regulation of the methionine cycle to sustain ESC pluripotency. Here, we show that adenosylhomocysteinase (AHCY), an important enzyme in the methionine cycle, is critical for the maintenance and differentiation of mouse embryonic stem cells (mESCs). We show that mESCs exhibit high levels of methionine metabolism, whereas decreasing methionine metabolism via depletion of AHCY promotes mESCs to differentiate into the three germ layers. AHCY is posttranslationally modified with an O-linked ß-N-acetylglucosamine sugar (O-GlcNAcylation), which is rapidly removed upon differentiation. O-GlcNAcylation of threonine 136 on AHCY increases its activity and is important for the maintenance of trimethylation of histone H3 lysine 4 (H3K4me3) to sustain mESC pluripotency. Blocking glycosylation of AHCY decreases the ratio of S-adenosylmethionine versus S-adenosylhomocysteine (SAM/SAH), reduces the level of H3K4me3, and poises mESC for differentiation. In addition, blocking glycosylation of AHCY reduces somatic cell reprogramming. Thus, our findings reveal a critical role of AHCY and a mechanistic understanding of O-glycosylation in regulating ESC pluripotency and differentiation.


Assuntos
Metionina/metabolismo , Células-Tronco Pluripotentes/metabolismo , Adenosil-Homocisteinase/metabolismo , Animais , Autorrenovação Celular , Reprogramação Celular , Glicosilação , Células HEK293 , Humanos , Camundongos , Células-Tronco Embrionárias Murinas/citologia , Células-Tronco Embrionárias Murinas/metabolismo , Células NIH 3T3
12.
Appl Environ Microbiol ; 86(9)2020 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-32111594

RESUMO

Lactococcus lactis encounters various environmental challenges, especially acid stress, during its growth. The cell wall can maintain the integrity and shape of the cell under environmental stress, and d-amino acids play an important role in cell wall synthesis. Here, by analyzing the effects of 19 different d-amino acids on the physiology of L. lactis F44, we found that exogenously supplied d-methionine and d-phenylalanine increased the nisin yield by 93.22% and 101.29%, respectively, as well as significantly increasing the acid resistance of L. lactis F44. The composition of the cell wall in L. lactis F44 with exogenously supplied d-Met or d-Phe was further investigated via a vancomycin fluorescence experiment and a liquid chromatography-mass spectrometry assay, which demonstrated that d-Met could be incorporated into the fifth position of peptidoglycan (PG) muropeptides and d-Phe could be added to the fourth and fifth positions. Moreover, overexpression of the PG synthesis gene murF further enhanced the levels of d-Met and d-Phe involved in PG and increased the survival rate under acid stress and the nisin yield of the strain. This study reveals that the exogenous supply of d-Met or d-Phe can change the composition of the cell wall and influence acid tolerance as well as nisin yield in L. lactis IMPORTANCE As d-amino acids play an important role in cell wall synthesis, we analyzed the effects of 19 different d-amino acids on L. lactis F44, demonstrating that d-Met and d-Phe can participate in peptidoglycan (PG) synthesis and improve the acid resistance and nisin yield of this strain. murF overexpression further increased the levels of d-Met and d-Phe incorporated into PG and contributed to the acid resistance of the strain. These findings suggest that d-Met and d-Phe can be incorporated into PG to improve the acid resistance and nisin yield of L. lactis, and this study provides new ideas for the enhancement of nisin production.


Assuntos
Ácidos/metabolismo , Parede Celular/fisiologia , Lactococcus lactis/metabolismo , Metionina/metabolismo , Nisina/biossíntese , Fenilalanina/metabolismo
13.
Life Sci ; 253: 117360, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32001269

RESUMO

AIMS: Progesterone receptor membrane component 1 (PGRMC1) has been reported to mediate the neuroprotective effect of progesterone, but the exact mechanism has not been elucidated. Therefore, the purpose of this study was to investigate the signalling pathway downstream of PGRMC1 in progesterone-induced neuroprotection. Recognition of the mechanism of progesterone opens novel perspectives for the treatment of diseases of the nervous system. MAIN METHODS: The PGRMC1 protein level was knocked down in rat primary cortical neurons, and Aß25-35 was used to establish an Alzheimer's disease cell model. The neuroprotective effect of progesterone was assessed by Hoechst 33258 staining and a cell counting kit-8 (CCK-8) assay. Then, proteomic and bioinformatic methods were used to analyse the proteins altered in response to PGRMC1 silencing to identify target proteins and signalling pathways involved in PGRMC1-mediated progesterone-induced neuroprotection. These findings were further verified by using signalling pathway inhibitors and western blotting. KEY FINDINGS: The neuroprotective effect of progesterone was significantly attenuated with PGRMC1 silencing. The expression of many proteins in the Ras signalling pathway was significantly changed in response to PGRMC1 silencing. FTI-277 inhibited progesterone-induced neuroprotection. Progesterone increased the expression of total Ras and Grb2. SIGNIFICANCE: These findings provide new perspectives for understanding the mechanism of and role of PGRMC1 in progesterone-induced neuroprotection. The Ras signalling pathway is the signalling pathway downstream of PGRMC1 in the mediation of progesterone-induced neuroprotection.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Peptídeos beta-Amiloides/química , Proteínas de Membrana/metabolismo , Neuroproteção/efeitos dos fármacos , Progesterona/metabolismo , Receptores de Progesterona/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Apoptose , Sobrevivência Celular , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Cromatografia Líquida de Alta Pressão , Inibidores Enzimáticos/química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Proteína Adaptadora GRB2/metabolismo , Técnicas de Inativação de Genes/métodos , Inativação Gênica , Humanos , Metionina/análogos & derivados , Metionina/química , Metionina/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Proteômica , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Espectrometria de Massas em Tandem
14.
Poult Sci ; 99(2): 1028-1037, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32036960

RESUMO

The research hypothesis postulated that the optimal dietary inclusion levels and ratios of lysine (Lys), arginine (Arg), and methionine (Met) can increase the growth potential of hybrid turkeys and limit metabolic disorders that weaken immune function. The experiment was carried out in a full rearing cycle, from 1 to 16 wk of age, in a two-factorial randomized design with 3 levels of Arg and 2 levels of Met (90, 100 and 110% of Arg, and 30 or 45% of Met, relative to the content of dietary Lys), with 6 groups of 8 replicates per group and 18 turkeys per replicate. In the first and second month of rearing, a significant dietary Arg-by-Met interaction was noted for daily feed intake and body weight gain, and a more beneficial effect was exerted by higher Met content and medium Arg content. Throughout the experiment, the higher dietary Met level increased the final body weight (BW) of turkeys (P = 0.001). Different dietary Arg levels had no influence on the growth performance of turkeys, but the lowest level decreased dressing yield (P = 0.001), and the highest level increased the percentage of breast muscles in the final BW of turkeys (P = 0.003). The lowest Arg level (90% of Lys content) undesirably increased the concentration of the proinflammatory cytokine IL-6 (P = 0.028) and decreased globulin concentration (P = 0.001) in the blood plasma of turkeys. The higher dietary Met level (45% of Lys content) increased plasma albumin concentration (P = 0.016). It can be concluded that higher dietary levels of Met (45 vs. 30% of Lys content) and Arg (100 and 110 vs. 90% of Lys content) have a more beneficial effect on the growth performance and immune status of turkeys.


Assuntos
Arginina/metabolismo , Imunidade Inata/efeitos dos fármacos , Lisina/metabolismo , Carne/análise , Metionina/metabolismo , Perus/fisiologia , Ração Animal/análise , Animais , Arginina/administração & dosagem , Dieta/veterinária , Suplementos Nutricionais/análise , Feminino , Lisina/administração & dosagem , Metionina/administração & dosagem , Perus/crescimento & desenvolvimento , Perus/imunologia
15.
J Med Chem ; 63(6): 2814-2832, 2020 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-32069402

RESUMO

Targeting polyamine metabolism is a proven anticancer strategy. Cancers often escape the polyamine biosynthesis inhibitors by increased polyamine import. Therefore, there is much interest in identifying polyamine transport inhibitors (PTIs) to be used in combination therapies. In a search for new PTIs, we serendipitously discovered a LAT-1 efflux agonist, which induces intracellular depletion of methionine, leucine, spermidine, and spermine, but not putrescine. Because S-adenosylmethioninamine is made from methionine, a loss of intracellular methionine leads to an inability to biosynthesize spermidine, and spermine. Importantly, we found that this methionine-depletion approach to polyamine depletion could not be rescued by exogenous polyamines, thereby obviating the need for a PTI. Using 3H-leucine (the gold standard for LAT-1 transport studies) and JPH-203 (a specific LAT-1 inhibitor), we showed that the efflux agonist did not inhibit the uptake of extracellular leucine but instead facilitated the efflux of intracellular leucine pools.


Assuntos
Transporte Biológico/efeitos dos fármacos , Transportador 1 de Aminoácidos Neutros Grandes/metabolismo , Poliaminas/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Benzoxazóis/farmacologia , Células CHO , Cricetulus , Descoberta de Drogas , Humanos , Leucina/metabolismo , Metionina/metabolismo , Putrescina/metabolismo , Bibliotecas de Moléculas Pequenas/química , Espermidina/metabolismo , Espermina/metabolismo , Tirosina/análogos & derivados , Tirosina/farmacologia
16.
Food Funct ; 11(2): 1764-1778, 2020 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-32044910

RESUMO

Dietary methionine restriction (MR) has been reported to extend lifespan, reduce obesity and decrease oxidative damage to mtDNA in the heart of rats, and increase endogenous hydrogen sulfide (H2S) production in the liver and blood. H2S has many potential benefits in the pathophysiology of the cardiovascular system. MR also increases the level of homocysteine (Hcy) in the liver and plasma, but elevated plasma Hcy is a risk factor for cardiovascular disease. Therefore, this study aimed to determine the effect of MR on cardiac function and metabolic status in obese middle-aged mice and its possible mechanisms. C57BL/6J mice (aged approximately 28 weeks) were divided into six dietary groups: CON (0.86% methionine + 4% fat), CMR40 (0.52% methionine + 4% fat), CMR80 (0.17% methionine + 4% fat), HFD (0.86% methionine + 24% fat), HMR40 (0.52% methionine + 24% fat) and HMR80 (0.17% methionine + 24% fat) for 15 consecutive weeks. Our results showed that 80% MR improves systolic dysfunction in middle-aged obese mice and enhances myocardial energy metabolism. 80% MR also reduces myocardial oxidative stress and improves inflammatory response. In addition, 80% MR increased mice Hcy levels and activated remethylation and transsulfur pathways of Hcy and promoted endogenous H2S production in the heart. 40% MR has the same trend, but is not significant. Moreover 40% MR at variance with 80% MR, did not decrease the body weight in both control and high-fat diet mice. These findings suggest that MR can improve myocardial energy metabolism, reduce heart inflammation and oxidative stress by increasing cardiac H2S production, and improve cardiac dysfunction in middle-aged obese mice.


Assuntos
Dieta , Metionina , Miocárdio , Obesidade/metabolismo , Animais , Peso Corporal , Cardiomegalia/metabolismo , Metabolismo Energético/fisiologia , Homocisteína/metabolismo , Sulfeto de Hidrogênio/metabolismo , Masculino , Metionina/administração & dosagem , Metionina/metabolismo , Metionina/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Miocárdio/citologia , Miocárdio/metabolismo , Estresse Oxidativo/fisiologia
17.
Obesity (Silver Spring) ; 28(3): 581-589, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32012481

RESUMO

OBJECTIVE: Restricting dietary methionine to 0.17% in male mice increases energy expenditure, reduces fat deposition, and improves metabolic health. The goal of this work was to compare each of these responses in postweaning male and female mice and in physically mature male and female mice. METHODS: Methionine-restricted (MR) diets were fed to age-matched cohorts of male and female mice for 8 to 10 weeks beginning at 8 weeks of age or beginning at 4 months of age. The physiological and transcriptional responses to MR were compared in the respective cohorts. RESULTS: Dietary MR produced sexually dimorphic changes in body composition in young growing animals, with males preserving lean at the expense of fat and females preserving fat at the expense of lean. The effects of MR on energy balance were comparable between sexes when the diet was initiated after attainment of physical maturity (4 months), and metabolic and endocrine responses were also comparable between males and females after 8 weeks on the MR diet. CONCLUSIONS: The sexually dimorphic effects of MR are limited to nutrient partitioning between lean and fat tissue deposition in young, growing mice. Introduction of the diet after physical maturity produced comparable effects on growth and metabolic responses in male and female mice.


Assuntos
Composição Corporal/efeitos dos fármacos , Dieta/efeitos adversos , Metabolismo Energético/efeitos dos fármacos , Metionina/efeitos adversos , Caracteres Sexuais , Fatores Etários , Animais , Humanos , Masculino , Metionina/metabolismo , Camundongos
18.
Diabetes ; 69(3): 465-476, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32029481

RESUMO

Children at increased genetic risk for type 1 diabetes (T1D) after environmental exposures may develop pancreatic islet autoantibodies (IA) at a very young age. Metabolic profile changes over time may imply responses to exposures and signal development of the first IA. Our present research in The Environmental Determinants of Diabetes in the Young (TEDDY) study aimed to identify metabolome-wide signals preceding the first IA against GAD (GADA-first) or against insulin (IAA-first). We profiled metabolomes by mass spectrometry from children's plasma at 3-month intervals after birth until appearance of the first IA. A trajectory analysis discovered each first IA preceded by reduced amino acid proline and branched-chain amino acids (BCAAs), respectively. With independent time point analysis following birth, we discovered dehydroascorbic acid (DHAA) contributing to the risk of each first IA, and γ-aminobutyric acid (GABAs) associated with the first autoantibody against insulin (IAA-first). Methionine and alanine, compounds produced in BCAA metabolism and fatty acids, also preceded IA at different time points. Unsaturated triglycerides and phosphatidylethanolamines decreased in abundance before appearance of either autoantibody. Our findings suggest that IAA-first and GADA-first are heralded by different patterns of DHAA, GABA, multiple amino acids, and fatty acids, which may be important to primary prevention of T1D.


Assuntos
Autoanticorpos/imunologia , Diabetes Mellitus Tipo 1/metabolismo , Metaboloma , Sintomas Prodrômicos , Alanina/metabolismo , Aminoácidos de Cadeia Ramificada , Pré-Escolar , Ácido Desidroascórbico/metabolismo , Diabetes Mellitus Tipo 1/imunologia , Ácidos Graxos/metabolismo , Feminino , Predisposição Genética para Doença , Glutamato Descarboxilase/imunologia , Humanos , Lactente , Recém-Nascido , Anticorpos Anti-Insulina/imunologia , Estudos Longitudinais , Masculino , Metionina/metabolismo , Fosfatidiletanolaminas/metabolismo , Prolina/metabolismo , Risco , Triglicerídeos/metabolismo , Ácido gama-Aminobutírico/metabolismo
19.
PLoS Comput Biol ; 16(1): e1007600, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31917825

RESUMO

Designed enzymes are of fundamental and technological interest. Experimental directed evolution still has significant limitations, and computational approaches are a complementary route. A designed enzyme should satisfy multiple criteria: stability, substrate binding, transition state binding. Such multi-objective design is computationally challenging. Two recent studies used adaptive importance sampling Monte Carlo to redesign proteins for ligand binding. By first flattening the energy landscape of the apo protein, they obtained positive design for the bound state and negative design for the unbound. We have now extended the method to design an enzyme for specific transition state binding, i.e., for its catalytic power. We considered methionyl-tRNA synthetase (MetRS), which attaches methionine (Met) to its cognate tRNA, establishing codon identity. Previously, MetRS and other synthetases have been redesigned by experimental directed evolution to accept noncanonical amino acids as substrates, leading to genetic code expansion. Here, we have redesigned MetRS computationally to bind several ligands: the Met analog azidonorleucine, methionyl-adenylate (MetAMP), and the activated ligands that form the transition state for MetAMP production. Enzyme mutants known to have azidonorleucine activity were recovered by the design calculations, and 17 mutants predicted to bind MetAMP were characterized experimentally and all found to be active. Mutants predicted to have low activation free energies for MetAMP production were found to be active and the predicted reaction rates agreed well with the experimental values. We suggest the present method should become the paradigm for computational enzyme design.


Assuntos
Enzimas , Método de Monte Carlo , Ligação Proteica/genética , Engenharia de Proteínas/métodos , Especificidade por Substrato/genética , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/química , Monofosfato de Adenosina/metabolismo , Azidas/química , Azidas/metabolismo , Sítios de Ligação/genética , Catálise , Enzimas/química , Enzimas/genética , Enzimas/metabolismo , Metionina/análogos & derivados , Metionina/química , Metionina/metabolismo , Metionina tRNA Ligase/química , Metionina tRNA Ligase/genética , Metionina tRNA Ligase/metabolismo , Mutação/genética , Norleucina/análogos & derivados , Norleucina/química , Norleucina/metabolismo
20.
Microbes Environ ; 35(1)2020.
Artigo em Inglês | MEDLINE | ID: mdl-31932540

RESUMO

Chitin amendment is an agricultural management strategy for controlling soil-borne plant disease. We previously reported an exponential decrease in chitin added to incubated upland soil. We herein investigated the transition of the bacterial community structure in chitin-degrading soil samples over time and the characteristics of chitinolytic bacteria in order to elucidate changes in the chitinolytic bacterial community structure during chitin degradation. The addition of chitin to soil immediately increased the population of bacteria in the genus Streptomyces, which is the main decomposer of chitin in soil environments. Lysobacter, Pseudoxanthomonas, Cellulosimicrobium, Streptosporangium, and Nonomuraea populations increased over time with decreases in that of Streptomyces. We isolated 104 strains of chitinolytic bacteria, among which six strains were classified as Lysobacter, from chitin-treated soils. These results suggested the involvement of Lysobacter as well as Streptomyces as chitin decomposers in the degradation of chitin added to soil. Lysobacter isolates required yeast extract or casamino acid for significant growth on minimal agar medium supplemented with glucose. Further nutritional analyses demonstrated that the six chitinolytic Lysobacter isolates required methionine (Met) to grow, but not cysteine or homocysteine, indicating Met auxotrophy. Met auxotrophy was also observed in two of the five type strains of Lysobacter spp. tested, and these Met auxotrophs used d-Met as well as l-Met. The addition of Met to incubated upland soil increased the population of Lysobacter. Met may be a factor increasing the population of Lysobacter in chitin-treated upland soil.


Assuntos
Bactérias/isolamento & purificação , Bactérias/metabolismo , Quitina/farmacologia , Metionina/metabolismo , Microbiota/efeitos dos fármacos , Microbiologia do Solo , Bactérias/classificação , Bactérias/genética , Quitina/análise , Quitina/metabolismo , Lysobacter/classificação , Lysobacter/genética , Lysobacter/isolamento & purificação , Lysobacter/metabolismo , Filogenia , RNA Ribossômico 16S/genética , Solo/química , Streptomyces/classificação , Streptomyces/genética , Streptomyces/isolamento & purificação , Streptomyces/metabolismo
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