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1.
Adv Exp Med Biol ; 1158: 59-70, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31452135

RESUMO

Mitochondria have a central role in cellular metabolism and reversible post-translational modifications regulate activity of mitochondrial proteins. Thanks to advances in proteomics, lysine acetylation has arisen as an important post-translational modification in the mitochondrion. During acetylation an acetyl group is covalently attached to the epsilon amino group in the side chain of lysine residues using acetyl-CoA as the substrate donor. Therefore the positive charge is neutralized, and this can affect the function of proteins thereby regulating enzyme activity, protein interactions, and protein stability. The major deacetylase in mitochondria is SIRT3 whose activity regulates many mitochondrial enzymes. The method of choice for the analysis of acetylated proteins foresees the combination of mass spectrometry-based proteomics with affinity enrichment techniques. Beyond the identification of lysine-acetylated proteins, many studies are moving towards the characterization of acetylated patterns in different diseases. Indeed, modifications in lysine acetylation status can directly alter mitochondrial function and, therefore, be linked to human diseases such as metabolic diseases, cancer, myocardial injury and neurodegenerative diseases. Despite the progress in the characterization of different lysine acetylation sites, additional studies are needed to differentiate the specific changes with a significant biological relevance.


Assuntos
Lisina , Mitocôndrias , Fenótipo , Acetilação , Humanos , Lisina/metabolismo , Mitocôndrias/química , Mitocôndrias/metabolismo , Proteínas Mitocondriais/química , Proteínas Mitocondriais/metabolismo , Processamento de Proteína Pós-Traducional
2.
Cancer Sci ; 110(10): 3145-3156, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31393050

RESUMO

Yes-associated protein (YAP) is a component of the canonical Hippo signaling pathway that is known to play essential roles in modulating organ size, development, and tumorigenesis. Activation or upregulation of YAP1, which contributes to cancer cell survival and chemoresistance, has been verified in different types of human cancers. However, the molecular mechanism of YAP1 upregulation in cancer is still unclear. Here we report that the E3 ubiquitin ligase STUB1 ubiquitinates and destabilizes YAP1, thereby inhibiting cancer cell survival. Low levels of STUB1 expression were correlated with increased protein levels of YAP1 in human gastric cancer cell lines and patient samples. Moreover, we revealed that STUB1 ubiquitinates YAP1 at the K280 site by K48-linked polyubiquitination, which in turn increases YAP1 turnover and promotes cellular chemosensitivity. Overall, our study establishes YAP1 ubiquitination and degradation mediated by the E3 ligase STUB1 as an important regulatory mechanism in gastric cancer, and provides a rationale for potential therapeutic interventions.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/química , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Resistencia a Medicamentos Antineoplásicos , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Neoplasias Gástricas/patologia , Ubiquitina-Proteína Ligases/metabolismo , Animais , Carcinogênese , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Células HEK293 , Humanos , Lisina/metabolismo , Camundongos , Transplante de Neoplasias , Estabilidade Proteica , Proteólise , Transdução de Sinais , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismo , Ubiquitinação
3.
Nat Commun ; 10(1): 2950, 2019 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-31270318

RESUMO

X-chromosome inactivation triggers fusion of A/B compartments to inactive X (Xi)-specific structures known as S1 and S2 compartments. SMCHD1 then merges S1/S2s to form the Xi super-structure. Here, we ask how S1/S2 compartments form and reveal that Xist RNA drives their formation via recruitment of Polycomb repressive complex 1 (PRC1). Ablating Smchd1 in post-XCI cells unveils S1/S2 structures. Loss of SMCHD1 leads to trapping Xist in the S1 compartment, impairing RNA spreading into S2. On the other hand, depleting Xist, PRC1, or HNRNPK precludes re-emergence of S1/S2 structures, and loss of S1/S2 compartments paradoxically strengthens the partition between Xi megadomains. Finally, Xi-reactivation in post-XCI cells can be enhanced by depleting both SMCHD1 and DNA methylation. We conclude that Xist, PRC1, and SMCHD1 collaborate in an obligatory, sequential manner to partition, fuse, and direct self-association of Xi compartments required for proper spreading of Xist RNA.


Assuntos
Proteínas Cromossômicas não Histona/metabolismo , Cromossomos de Mamíferos/genética , Complexo Repressor Polycomb 1/metabolismo , RNA Longo não Codificante/metabolismo , Cromossomo X/química , Cromossomo X/genética , Animais , Metilação de DNA/genética , Histonas/metabolismo , Lisina/metabolismo , Camundongos , Modelos Genéticos , Inativação do Cromossomo X/genética
4.
Nat Commun ; 10(1): 2909, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266957

RESUMO

Cells form and use biomolecular condensates to execute biochemical reactions. The molecular properties of non-membrane-bound condensates are directly connected to the amino acid content of disordered protein regions. Lysine plays an important role in cellular function, but little is known about its role in biomolecular condensation. Here we show that protein disorder is abundant in protein/RNA granules and lysine is enriched in disordered regions of proteins in P-bodies compared to the entire human disordered proteome. Lysine-rich polypeptides phase separate into lysine/RNA-coacervates that are more dynamic and differ at the molecular level from arginine/RNA-coacervates. Consistent with the ability of lysine to drive phase separation, lysine-rich variants of the Alzheimer's disease-linked protein tau undergo coacervation with RNA in vitro and bind to stress granules in cells. Acetylation of lysine reverses liquid-liquid phase separation and reduces colocalization of tau with stress granules. Our study establishes lysine as an important regulator of cellular condensation.


Assuntos
Lisina/metabolismo , RNA/química , RNA/metabolismo , Proteínas tau/química , Proteínas tau/metabolismo , Acetilação , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Linhagem Celular , Grânulos Citoplasmáticos/genética , Grânulos Citoplasmáticos/metabolismo , Humanos , Lisina/química , Lisina/genética , RNA/genética , Proteínas tau/genética
5.
Food Chem ; 299: 125166, 2019 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-31323443

RESUMO

Dynamic high-pressure microfluidization (DHPM) pretreatment and glycation with lactose were employed to modify α-Lactalbumin (α-LA) with respect to the IgE/IgG binding capacities. No significant difference on incorporation ratio value of glycated α-LA was observed with and without DHPM pretreatment. However, IgE/IgG binding capacities of α-LA were decreased after glycation and DHPM pretreatment promoted the reduction. The lowest IgE/IgG binding capacities of glycated α-LA were obtained by DHPM pretreatment at 110 MPa. Native α-LA was mainly glycated at K62, K94, K98, whereas glycation sites and degree of substitution per peptide (DSP) were added after DHPM treatment. Therefore, the reduced IgE/IgG binding capacities of α-LA was attributed to the characteristics of glycated sites, including the amount, location, and DSP values. Interestingly, K98 played the most important role in decreasing IgE/IgG binding capacities of α-LA. The study revealed that glycation combined with DHPM was a promising way to decrease the allergenicity of proteins.


Assuntos
Hipersensibilidade Alimentar/imunologia , Imunoglobulina E/metabolismo , Imunoglobulina G/metabolismo , Lactalbumina/metabolismo , Alérgenos/química , Animais , Bovinos , Ensaio de Imunoadsorção Enzimática , Indústria de Processamento de Alimentos/métodos , Humanos , Lactalbumina/química , Lactalbumina/imunologia , Lactose/química , Lisina/metabolismo , Espectrometria de Massas/métodos , Pressão , Coelhos
6.
Immunogenetics ; 71(7): 489-499, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31297569

RESUMO

Epigenetic modifications have been shown to be important for immune cell differentiation by regulating gene transcription. However, the role and mechanism of histone methylation in the development and differentiation of iNKT cells in rheumatoid arthritis (RA) mice have yet to be deciphered. The DBA/1 mouse RA model was established by using a modified GPI mixed peptide. We demonstrated that total peripheral blood, thymus, and spleen iNKT cells in RA mice decreased significantly, while iNKT1 in the thymus and spleen was increased significantly. PLZF protein and PLZF mRNA levels were significantly decreased in thymus DP T cells, while T-bet protein and mRNA were significantly increased in thymus iNKT cells. We found a marked accumulation in H3K27me3 around the promoter regions of the signature gene Zbtb16 in RA mice thymus DP T cells, and an accumulation of H3K4me3 around the promoters of the Tbx21 gene in iNKT cells. The expression levels of UTX in the thymus of RA mice were significantly reduced. The changes in the above indicators were particularly significant in the progressive phase of inflammation (11 days after modeling) and the peak phase of inflammation (14 days after modeling) in RA mice. Developmental and differentiation defects of iNKT cells in RA mice were associated with abnormal methylation levels (H3K27me3 and H3K4me3) in the promoters of key genes Zbtb16 (encoding PLZF) and Tbx21 (encoding T-bet). Decreased UTX of thymus histone demethylase levels resulted in the accumulation of H3K27me3 modification.


Assuntos
Artrite Reumatoide/patologia , Lisina/metabolismo , Células T Matadoras Naturais/patologia , Regiões Promotoras Genéticas , Timo/fisiologia , Animais , Artrite Experimental/patologia , Diferenciação Celular , Epigênese Genética , Regulação da Expressão Gênica , Histona Desmetilases/genética , Histona Desmetilases/metabolismo , Metilação , Camundongos Endogâmicos DBA , Proteína com Dedos de Zinco da Leucemia Promielocítica/genética , Proteína com Dedos de Zinco da Leucemia Promielocítica/metabolismo , Baço/patologia , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo
7.
Food Chem ; 293: 396-407, 2019 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-31151627

RESUMO

To explore the involvement of protein lysine acetylation in the conversion of muscle to meat, a quantitative analysis of the acetylome in postmortem porcine muscle with or without antemortem stress was conducted. In total, 771 acetylpeptides containing 681 lysine acetylation sites mapping to 176 acetylproteins were identified. Acetylproteins were enriched in muscle contraction, carbohydrate metabolism, cell apoptosis and calcium signaling. Bioinformatic analysis suggests that preslaughter handling may be associated with glycolysis in postmortem muscle and the overall meat quality, via acetylation of multiple enzymes of glycogenolysis/glycolysis, regulate rigor mortis via acetylation of contractile, ATP production and calcium signaling-related proteins, and regulate stress response, cell apoptosis and meat tenderization via regulating the functions of heat shock proteins and permeability transition pore complex. This study provides the first overview of the acetylome in postmortem muscle as affected by preslaughter handling and broadens knowledge of the biochemistry regulating meat quality development.


Assuntos
Qualidade dos Alimentos , Lisina/metabolismo , Músculo Esquelético/metabolismo , Proteômica/métodos , Carne Vermelha/análise , Acetilação , Animais , Biologia Computacional/métodos , Glicólise , Proteínas de Choque Térmico/metabolismo , Proteínas de Carne/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Mudanças Depois da Morte , Estresse Psicológico , Suínos
8.
Mol Biol (Mosk) ; 53(3): 524-528, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31184618

RESUMO

Proteins with homo-repeats of more than 4 amino acid residues in length were examined to understand whether some splicing sites in pre-mRNA may be attributed to homo-repeats in human proteins. The human proteome was found to contain a total of 404 proteins with homo-repeats that account for at least one splicing site in pre-mRNA. Pre-mRNA splicing sites were more often found in the C-terminal part (67%) than in the middle orN-terminal part of a homo-repeat. Ten homo-repeats were identified to have two splicing sites per repeat. The repeats were lysine homo-repeats in all but one case.


Assuntos
Proteínas/análise , Proteínas/química , Precursores de RNA/genética , Sítios de Splice de RNA/genética , Sequências Repetitivas de Aminoácidos/genética , Humanos , Lisina/genética , Lisina/metabolismo , Proteínas/genética , Proteoma/análise , Proteoma/química , Proteoma/genética , Processamento de RNA/genética
9.
Microb Cell Fact ; 18(1): 106, 2019 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-31186003

RESUMO

BACKGROUND: Late-stage fermentation broth contains high concentrations of target chemicals. Additionally, it contains various cellular metabolites which have leaked from lysed cells, which would exert multifactorial stress to industrial hyperproducers and perturb both cellular metabolism and product formation. Although adaptive laboratory evolution (ALE) has been wildly used to improve stress tolerance of microbial cell factories, single-factor stress condition (i.e. target product or sodium chloride at a high concentration) is currently provided. In order to enhance bacterial stress tolerance to actual industrial production conditions, ALE in late-stage fermentation broth is desired. Genome replication engineering assisted continuous evolution (GREACE) employs mutants of the proofreading element of DNA polymerase complex (DnaQ) to facilitate mutagenesis. Application of GREACE coupled-with selection under stress conditions is expected to accelerate the ALE process. RESULTS: In this study, GREACE was first modified by expressing a DnaQ mutant KR5-2 using an arabinose inducible promoter on a temperature-sensitive plasmid, which resulted in timed mutagenesis introduction. Using this method, tolerance of a lysine hyperproducer E. coli MU-1 was improved by enriching mutants in a lysine endpoint fermentation broth. Afterwards, the KR5-2 expressing plasmid was cured to stabilize acquired genotypes. By subsequent fermentation evaluation, a mutant RS3 with significantly improved lysine production capacity was selected. The final titer, yield and total amount of lysine produced by RS3 in a 5-L batch fermentation reached 155.0 ± 1.4 g/L, 0.59 ± 0.02 g lysine/g glucose, and 605.6 ± 23.5 g, with improvements of 14.8%, 9.3%, and 16.7%, respectively. Further metabolomics and genomics analyses, coupled with molecular biology studies revealed that mutations SpeBA302V, AtpBS165N and SecYM145V mainly contributed both to improved cell integrity under stress conditions and enhanced metabolic flux into lysine synthesis. CONCLUSIONS: Our present study indicates that improving a lysine hyperproducer by GREACE-assisted ALE in its stressful living environment is efficient and effective. Accordingly, this is a promising method for improving other valuable chemical hyperproducers.


Assuntos
Evolução Molecular Direcionada/métodos , Escherichia coli/metabolismo , Lisina/metabolismo , Engenharia Metabólica/métodos , Escherichia coli/genética , Escherichia coli/crescimento & desenvolvimento , Fermentação , Mutagênese
10.
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
11.
Cancer Sci ; 110(8): 2408-2420, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31215094

RESUMO

Esophageal squamous cell carcinomas (ESCCs) as well as adenocarcinomas (EACs) were developed in rat duodenal contents reflux models (reflux model). The present study aimed to shed light on the mechanism by which bile acid stimulation causes cancer onset and progression. Metabolomics analyses were performed on samples of neoplastic and nonneoplastic tissues from reflux models, and K14D, cultivated from a nonmetastatic, primary ESCC, and ESCC-DR, established from a metastatic thoracic lesion. ESCC-DRtca2M was prepared by treating ESCC-DR cells with taurocholic acid (TCA) to accelerate cancer progression. The lines were subjected to comprehensive genomic analyses. In addition, protein expression levels of glucose-6-phosphate dehydrogenase (G6PD), nuclear factor kappa B (NF-κB) (p65) and O-linked N-Acetylglucosamine (O-GlcNAc) were compared among lines. Cancers developed in the reflux models exhibited greater hexosamine biosynthesis pathway (HBP) activation compared with the nonneoplastic tissues. Expression of O-GlcNAc transferase (OGT) increased considerably in both ESCC and EAC compared with nonneoplastic squamous epithelium. Conversely, cell line-based experiments revealed the greater activation of the pentose phosphate pathway (PPP) at higher degrees of malignancy. G6PD overexpression in response to TCA exposure was observed. Both NF-κB (p65) and O-GlcNAc were expressed more highly in ESCC-DRtca2M than in the other cell lines. Moreover, ESCC-DRtca2M cells had additional chromosomal abnormalities in excess of ESCC-DR cells. Overall, glucose metabolism was upregulated in both esophageal cancer tissue and cell lines. While bile acids are not mutagenic, chronic exposure seems to trigger NF-κB(p65) activation, potentially inducing genetic mutations as well as facilitating carcinogenesis and cancer progression. Glucose metabolism was upregulated in both esophageal cancer tissue and cell lines, and the HBP was activated in the former. The cell line-based experiments demonstrated upregulation of the pentose phosphate pathway (PPP) at higher degrees of malignancy. While bile acids are not mutagenic, chronic exposure seems to trigger G6PD overexpression and NF-κB (p65) activation, potentially inducing genetic mutations as well as facilitating carcinogenesis and cancer progression.


Assuntos
Ácidos e Sais Biliares/metabolismo , Vias Biossintéticas/fisiologia , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas do Esôfago/metabolismo , Hexosaminas/metabolismo , Via de Pentose Fosfato/fisiologia , Acetilglucosamina/metabolismo , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Linhagem Celular Tumoral , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/patologia , Regulação Neoplásica da Expressão Gênica/fisiologia , Glucose/análogos & derivados , Glucose/metabolismo , Glucosefosfato Desidrogenase/metabolismo , Lisina/análogos & derivados , Lisina/metabolismo , Masculino , NF-kappa B/metabolismo , Ratos , Ratos Wistar , Transdução de Sinais/fisiologia , Regulação para Cima/fisiologia
12.
Anim Sci J ; 90(8): 932-938, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31218788

RESUMO

Three rumen-protected lysine (RPL) products (AjiPro® -L, LysiPEARLTM , and Feedtech Bypass LysineTM : A, B, and C, respectively) were tested for stability in two forage-based total mixed rations (TMR1, 41.3% dry matter (DM), and TMR2, 49.5% DM) (experiment 1) and for Brix value (experiment 2). In experiment 1, each RPL product (2 g each) and TMR diet (200 g) were mixed and stored in plastic bags at 20°C for 0, 1, 3, 6, 12, 24, and 48 hr. In experiment 2, each RPL product (2 g) was dispensed into ion-exchanged water (20 ml) and kept at 20°C for 0, 0.5, 1, 3, 6, 12, 24, and 48 hr. At each time point, free lysine (Lys) content and Brix values of extracts were measured, and Lys release (LR, %) was calculated. All RPL products LR% varied with varying diets DM and increased with increasing of time exposed to diets; it was highest in C, followed by B, and then A. Water LR% positively correlated with that from diets and with Brix values of Lys dissociated in water. Our results indicated that Lys dissociation from RPL products is affected by diet DM content. Brix value may be used as a potential marker for RPL protection efficacy.


Assuntos
Ração Animal , Dieta/veterinária , Lisina/metabolismo , Rúmen/metabolismo , Animais , Técnicas In Vitro , Temperatura Ambiente , Fatores de Tempo , Água
13.
Nat Commun ; 10(1): 2399, 2019 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-31160585

RESUMO

Manganese superoxide dismutase (MnSOD) functions as a tumor suppressor; however, once tumorigenesis occurs, clinical data suggest MnSOD levels correlate with more aggressive human tumors, implying a potential dual function of MnSOD in the regulation of metabolism. Here we show, using in vitro transformation and xenograft growth assays that the MnSOD-K68 acetylation (Ac) mimic mutant (MnSODK68Q) functions as a tumor promoter. Interestingly, in various breast cancer and primary cell types the expression of MnSODK68Q is accompanied with a change of MnSOD's stoichiometry from a known homotetramer complex to a monomeric form. Biochemical experiments using the MnSOD-K68Q Ac-mimic, or physically K68-Ac (MnSOD-K68-Ac), suggest that these monomers function as a peroxidase, distinct from the established MnSOD superoxide dismutase activity. MnSODK68Q expressing cells exhibit resistance to tamoxifen (Tam) and cells selected for Tam resistance exhibited increased K68-Ac and monomeric MnSOD. These results suggest a MnSOD-K68-Ac metabolic pathway for Tam resistance, carcinogenesis and tumor progression.


Assuntos
Neoplasias da Mama/genética , Carcinogênese/genética , Resistencia a Medicamentos Antineoplásicos/genética , Superóxido Dismutase/genética , Acetilação , Animais , Antineoplásicos Hormonais/uso terapêutico , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Progressão da Doença , Humanos , Técnicas In Vitro , Lisina/metabolismo , Células MCF-7 , Camundongos , Mutação , Transplante de Neoplasias , Peroxidase/metabolismo , Estrutura Quaternária de Proteína/genética , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Tamoxifeno/uso terapêutico , Proteínas Supressoras de Tumor
14.
Cell Mol Life Sci ; 76(15): 2873-2883, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31123776

RESUMO

Lysine methylation, catalyzed by protein lysine methyltransferases (PKMTs), is a central post-translational modification regulating many signaling pathways. It has direct and indirect effects on chromatin structure and transcription. Accumulating evidence suggests that dysregulation of PKMT activity has a fundamental impact on the development of many pathologies. While most of these works involve in-depth analysis of methylation events in the context of histones, in recent years, it has become evident that methylation of non-histone proteins also plays a pivotal role in cell processes. This review highlights the importance of non-histone methylation, with focus on methylation events taking place in the nucleus. Known experimental platforms which were developed to identify new methylation events, as well as examples of specific lysine methylation signaling events which regulate key transcription factors, are presented. In addition, the role of these methylation events in normal and disease states is emphasized.


Assuntos
Núcleo Celular/metabolismo , Lisina/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , Humanos , Metilação , Análise Serial de Proteínas , Transdução de Sinais , Fatores de Transcrição/metabolismo
15.
J Anim Sci ; 97(7): 2822-2836, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31115459

RESUMO

Nutrient requirements of sows during lactation are related mainly to their milk yield and feed intake, and vary greatly among individuals. In practice, nutrient requirements are generally determined at the population level based on average performance. The objective of the present modeling approach was to explore the variability in nutrient requirements among sows by combining current knowledge about nutrient use with on-farm data available on sows at farrowing [parity, BW, backfat thickness (BT)] and their individual performance (litter size, litter average daily gain, daily sow feed intake) to estimate nutrient requirements. The approach was tested on a database of 1,450 lactations from 2 farms. The effects of farm (A, B), week of lactation (W1: week 1, W2: week 2, W3+: week 3 and beyond), and parity (P1: 1, P2: 2, P3+: 3 and beyond) on sow performance and their nutrient requirements were evaluated. The mean daily ME requirement was strongly correlated with litter growth (R2 = 0.95; P < 0.001) and varied slightly according to sow BW, which influenced the maintenance cost. The mean daily standardized ileal digestible (SID) lysine requirement was influenced by farm, week of lactation, and parity. Variability in SID lysine requirement per kg feed was related mainly to feed intake (R2 = 0.51; P < 0.001) and, to a smaller extent, litter growth (R2 = 0.27; P < 0.001). It was lowest in W1 (7.0 g/kg), greatest in W2 (7.9 g/kg), and intermediate in W3+ (7.5 g/kg; P < 0.001) because milk production increased faster than feed intake capacity did. It was lower for P3+ (6.7 g/kg) and P2 sows (7.3 g/kg) than P1 sows (8.3 g/kg) due to the greater feed intake of multiparous sows. The SID lysine requirement per kg of feed was met for 80% of sows when supplies were 112 and 120% of the mean population requirement on farm A and B, respectively, indicating higher variability in requirements on farm B. Other amino acid and mineral requirements were influenced in the same way as SID lysine. The present modeling approach allows to capture individual variability in the performance of sows and litters according to farm, stage of lactation, and parity. It is an initial step in the development of new types of models able to process historical farm data (e.g., for ex post assessment of nutrient requirements) and real-time data (e.g., to control precision feeding).


Assuntos
Aminoácidos/metabolismo , Ingestão de Alimentos , Ingestão de Energia , Leite/metabolismo , Minerais/metabolismo , Suínos/fisiologia , Animais , Feminino , Íleo/metabolismo , Lactação , Tamanho da Ninhada de Vivíparos , Lisina/metabolismo , Nutrientes/metabolismo , Necessidades Nutricionais , Paridade , Gravidez
16.
Int J Mol Sci ; 20(10)2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31091743

RESUMO

As one of the typical Maillard reaction products, furosine has been widely reported in a variety of heat-processed food. Though furosine was shown to be toxic on organs, its toxicity mechanism is still unclear. The present study aimed to investigate the toxicity mechanism of furosine in liver tissue. An intragastric gavage mice model (42-day administration, 0.1/0.25/0.5 g/kg of furosine per day) and a mice primary hepatocyte model were employed to investigate the toxicity mechanism of furosine on mice liver tissue. A metabonomics analysis of mice liver, serum, and red blood cells (RBC) was performed. The special metabolic mediator of furosine, lysophosphatidylcholine 18:0 (LPC (18:0)) was identified. Then, the effect of the upstream gene phospholipase A2 gamma (PLA2-3) on LPC (18:0), as well as the effect of furosine (100 mg/L) on the receptor-interacting serine/threonine-protein kinase (RIPK)1/RIPK3/mixed lineage kinase domain-like protein (MLKL) pathway and inflammatory factors, was determined in liver tissue and primary hepatocytes. PLA2-3 was found to regulate the level of LPC (18:0) and activate the expression of RIPK1, RIPK3, P-MLKL, and of the inflammatory factors including tumor necrosis factor α (TNF-α) and interleukin (IL-1ß), both in liver tissue and in primary hepatocytes. Upon treatment with furosine, the upstream sensor PLA2-3 activated the RIPK1/RIPK3/MLKL necroptosis pathway and caused inflammation by regulating the expression of LPC (18:0), which further caused liver damage.


Assuntos
Produtos Finais de Glicação Avançada/toxicidade , Hepatócitos/metabolismo , Lisina/análogos & derivados , Proteínas Quinases/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Morte Celular , Células Cultivadas , Feminino , Produtos Finais de Glicação Avançada/metabolismo , Interleucina-1beta/metabolismo , Lisina/metabolismo , Lisina/toxicidade , Masculino , Camundongos , Camundongos Endogâmicos ICR , Fator de Necrose Tumoral alfa/metabolismo
17.
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
18.
MBio ; 10(3)2019 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-31064836

RESUMO

Despite intensive study for 50 years, the biochemical and genetic links between lysine metabolism and central metabolism in Pseudomonas putida remain unresolved. To establish these biochemical links, we leveraged random barcode transposon sequencing (RB-TnSeq), a genome-wide assay measuring the fitness of thousands of genes in parallel, to identify multiple novel enzymes in both l- and d-lysine metabolism. We first describe three pathway enzymes that catabolize l-2-aminoadipate (l-2AA) to 2-ketoglutarate (2KG), connecting d-lysine to the TCA cycle. One of these enzymes, P. putida 5260 (PP_5260), contains a DUF1338 domain, representing a family with no previously described biological function. Our work also identified the recently described coenzyme A (CoA)-independent route of l-lysine degradation that results in metabolization to succinate. We expanded on previous findings by demonstrating that glutarate hydroxylase CsiD is promiscuous in its 2-oxoacid selectivity. Proteomics of selected pathway enzymes revealed that expression of catabolic genes is highly sensitive to the presence of particular pathway metabolites, implying intensive local and global regulation. This work demonstrated the utility of RB-TnSeq for discovering novel metabolic pathways in even well-studied bacteria, as well as its utility a powerful tool for validating previous research.IMPORTANCE P. putida lysine metabolism can produce multiple commodity chemicals, conferring great biotechnological value. Despite much research, the connection of lysine catabolism to central metabolism in P. putida remained undefined. Here, we used random barcode transposon sequencing to fill the gaps of lysine metabolism in P. putida We describe a route of 2-oxoadipate (2OA) catabolism, which utilizes DUF1338-containing protein P. putida 5260 (PP_5260) in bacteria. Despite its prevalence in many domains of life, DUF1338-containing proteins have had no known biochemical function. We demonstrate that PP_5260 is a metalloenzyme which catalyzes an unusual route of decarboxylation of 2OA to d-2-hydroxyglutarate (d-2HG). Our screen also identified a recently described novel glutarate metabolic pathway. We validate previous results and expand the understanding of glutarate hydroxylase CsiD by showing that can it use either 2OA or 2KG as a cosubstrate. Our work demonstrated that biological novelty can be rapidly identified using unbiased experimental genetics and that RB-TnSeq can be used to rapidly validate previous results.


Assuntos
Aptidão Genética , Lisina/metabolismo , Pseudomonas putida/enzimologia , Pseudomonas putida/genética , Redes e Vias Metabólicas
19.
J Agric Food Chem ; 67(23): 6594-6602, 2019 Jun 12.
Artigo em Inglês | MEDLINE | ID: mdl-31091091

RESUMO

Modifications of lysine contribute to the amount of dietary advanced glycation end-products reaching the colon. However, little is known about the ability of intestinal bacteria to metabolize dietary N-ε-carboxymethyllysine (CML). Successive transfers of fecal microbiota in growth media containing CML were used to identify and isolate species able to metabolize CML under anaerobic conditions. From our study, only donors exposed to processed foods degraded CML, and anaerobic bacteria enrichments from two of them used 77 and 100% of CML. Oscillibacter and Cloacibacillus evryensis increased in the two donors after the second transfer, highlighting that the bacteria from these taxa could be candidates for anaerobic CML degradation. A tentative identification of CML metabolites produced by a pure culture of Cloacibacillus evryensis was performed by mass spectrometry: carboxymethylated biogenic amines and carboxylic acids were identified as CML degradation products. The study confirmed the ability of intestinal bacteria to metabolize CML under anoxic conditions.


Assuntos
Bactérias/metabolismo , Colo/microbiologia , Microbioma Gastrointestinal , Produtos Finais de Glicação Avançada/metabolismo , Lisina/análogos & derivados , Adulto , Anaerobiose , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Fezes/microbiologia , Produtos Finais de Glicação Avançada/química , Glicosilação , Humanos , Lactente , Lisina/química , Lisina/metabolismo
20.
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
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