RESUMO
The human microbiome exists throughout the body, and it is essential for maintaining various physiological processes, including immunity, and dysbiotic events, which are associated with autoimmunity. Peptidylarginine deiminase (PAD) enzymes can citrullinate self-proteins related to rheumatoid arthritis (RA) that induce the production of anti-citrullinated protein antibodies (ACPAs) and lead to inflammation and joint damage. The present investigation was carried out to demonstrate the expression of homologs of PADs or arginine deiminases (ADs) and citrullinated proteins in members of the human microbiota. To achieve the objective, we used 17 microbial strains and specific polyclonal antibodies (pAbs) of the synthetic peptide derived from residues 100-200 of human PAD2 (anti-PAD2 pAb), and the recombinant fragment of amino acids 326 and 611 of human PAD4 (anti-PAD4 pAb), a human anti-citrulline pAb, and affinity ACPAs of an RA patient. Western blot (WB), enzyme-linked immunosorbent assay (ELISA), elution, and a test with Griess reagent were used. This is a cross-sectional case-control study on patients diagnosed with RA and control subjects. Inferential statistics were applied using the non-parametric Kruskal-Wallis test and Mann-Whitney U test generated in the SPSS program. Some members of phyla Firmicutes and Proteobacteria harbor homologs of PADs/ADs and citrullinated antigens that are reactive to the ACPAs of RA patients. Microbial citrullinome and homolog enzymes of PADs/ADs are extensive in the human microbiome and are involved in the production of ACPAs. Our findings suggest a molecular link between microorganisms of a dysbiotic microbiota and RA pathogenesis.
Assuntos
Anticorpos Antiproteína Citrulinada , Artrite Reumatoide , Citrulinação , Microbiota , Desiminases de Arginina em Proteínas , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Anticorpos Antiproteína Citrulinada/imunologia , Anticorpos Antiproteína Citrulinada/metabolismo , Artrite Reumatoide/imunologia , Artrite Reumatoide/microbiologia , Estudos de Casos e Controles , Citrulina/metabolismo , Estudos Transversais , Hidrolases/metabolismo , Proteína-Arginina Desiminase do Tipo 2/metabolismo , Proteína-Arginina Desiminase do Tipo 4/metabolismo , Desiminases de Arginina em Proteínas/metabolismo , Desiminases de Arginina em Proteínas/genéticaRESUMO
Identification of genetic modulators of lysosomal enzyme activities and glycosphingolipids (GSLs) may facilitate the development of therapeutics for diseases in which they participate, including Lysosomal Storage Disorders (LSDs). To this end, we used a systems genetics approach: we measured 11 hepatic lysosomal enzymes and many of their natural substrates (GSLs), followed by modifier gene mapping by GWAS and transcriptomics associations in a panel of inbred strains. Unexpectedly, most GSLs showed no association between their levels and the enzyme activity that catabolizes them. Genomic mapping identified 30 shared predicted modifier genes between the enzymes and GSLs, which are clustered in three pathways and are associated with other diseases. Surprisingly, they are regulated by ten common transcription factors, and their majority by miRNA-340p. In conclusion, we have identified novel regulators of GSL metabolism, which may serve as therapeutic targets for LSDs and may suggest the involvement of GSL metabolism in other pathologies.
Assuntos
Glicoesfingolipídeos , Doenças por Armazenamento dos Lisossomos , Animais , Camundongos , Glicoesfingolipídeos/metabolismo , Doenças por Armazenamento dos Lisossomos/metabolismo , Hidrolases/metabolismo , Lisossomos/metabolismoRESUMO
Dysfunction of the adipose tissue metabolism is considered as a significant hallmark of aging. It has been proposed that α-ß hydrolase domain containing 5 (ABHD5) plays a critical role in the control of lipolysis. However, the role of ABHD5 in the control of lipolysis during aging or exercise is unknown. Here we combined the experimental mouse model with transcriptomic analyzes by using murine and human databases to explore the role of ABHD5 in the adipose tissue during aging and in response to exercise. Transcriptomic data revealed a downregulation of Abhd5 messenger RNA levels in the subcutaneous white adipose tissue (scWAT) over time in individuals from 20 to 69 years old. Aged mice displayed dramatic reduction of ABHD5 protein content and lipolytic-related proteins in the scWAT. Interestingly, 4 weeks of high-intensity interval training increased ABHD5 protein level and restored the lipolytic pathway in the scWAT of aged mice. Altogether, our findings demonstrated that aging affects ABHD5 content in the adipose tissue of mice and humans. Conversely, exercise increases ABHD5 activity, recovering the lipolytic activity in aged mice.
Assuntos
1-Acilglicerol-3-Fosfato O-Aciltransferase , Tecido Adiposo , Envelhecimento , Exercício Físico , Lipólise , Adulto , Idoso , Animais , Humanos , Camundongos , Pessoa de Meia-Idade , Adulto Jovem , 1-Acilglicerol-3-Fosfato O-Aciltransferase/genética , 1-Acilglicerol-3-Fosfato O-Aciltransferase/metabolismo , Tecido Adiposo/enzimologia , Envelhecimento/metabolismo , Hidrolases/genética , Hidrolases/metabolismoRESUMO
Cell surface receptors play essential roles in perceiving and processing external and internal signals at the cell surface of plants and animals. The receptor-like protein kinases (RLK) and receptor-like proteins (RLPs), two major classes of proteins with membrane receptor configuration, play a crucial role in plant development and disease defense. Although RLPs and RLKs share a similar single-pass transmembrane configuration, RLPs harbor short divergent C-terminal regions instead of the conserved kinase domain of RLKs. This RLP receptor structural design precludes sequence comparison algorithms from being used for high-throughput predictions of the RLP family in plant genomes, as has been extensively performed for RLK superfamily predictions. Here, we developed the RLPredictiOme, implemented with machine learning models in combination with Bayesian inference, capable of predicting RLP subfamilies in plant genomes. The ML models were simultaneously trained using six types of features, along with three stages to distinguish RLPs from non-RLPs (NRLPs), RLPs from RLKs, and classify new subfamilies of RLPs in plants. The ML models achieved high accuracy, precision, sensitivity, and specificity for predicting RLPs with relatively high probability ranging from 0.79 to 0.99. The prediction of the method was assessed with three datasets, two of which contained leucine-rich repeats (LRR)-RLPs from Arabidopsis and rice, and the last one consisted of the complete set of previously described Arabidopsis RLPs. In these validation tests, more than 90% of known RLPs were correctly predicted via RLPredictiOme. In addition to predicting previously characterized RLPs, RLPredictiOme uncovered new RLP subfamilies in the Arabidopsis genome. These include probable lipid transfer (PLT)-RLP, plastocyanin-like-RLP, ring finger-RLP, glycosyl-hydrolase-RLP, and glycerophosphoryldiester phosphodiesterase (GDPD, GDPDL)-RLP subfamilies, yet to be characterized. Compared to the only Arabidopsis GDPDL-RLK, molecular evolution studies confirmed that the ectodomain of GDPDL-RLPs might have undergone a purifying selection with a predominance of synonymous substitutions. Expression analyses revealed that predicted GDPGL-RLPs display a basal expression level and respond to developmental and biotic signals. The results of these biological assays indicate that these subfamily members have maintained functional domains during evolution and may play relevant roles in development and plant defense. Therefore, RLPredictiOme provides a framework for genome-wide surveys of the RLP superfamily as a foundation to rationalize functional studies of surface receptors and their relationships with different biological processes.
Assuntos
Arabidopsis , Proteínas de Plantas , Animais , Proteínas de Plantas/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Plastocianina/genética , Plastocianina/metabolismo , Teorema de Bayes , Leucina/metabolismo , Plantas/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Receptores de Superfície Celular/metabolismo , Aprendizado de Máquina , Hidrolases/metabolismo , Diester Fosfórico Hidrolases/metabolismo , Lipídeos , FilogeniaRESUMO
Amyloids are supramolecular assemblies composed of polypeptides stabilized by an intermolecular beta-sheet core. These misfolded conformations have been traditionally associated with pathological conditions such as Alzheimer's and Parkinson´s diseases. However, this classical paradigm has changed in the last decade since the discovery that the amyloid state represents a universal alternative fold accessible to virtually any polypeptide chain. Moreover, recent findings have demonstrated that the amyloid fold can serve as catalytic scaffolds, creating new opportunities for the design of novel active bionanomaterials. Here, we review the latest advances in this area, with particular emphasis on the design and development of catalytic amyloids that exhibit hydrolytic activities. To date, three different types of activities have been demonstrated: esterase, phosphoesterase and di-phosphohydrolase. These artificial hydrolases emerge upon the self-assembly of small peptides into amyloids, giving rise to catalytically active surfaces. The highly stable nature of the amyloid fold can provide an attractive alternative for the design of future synthetic hydrolases with diverse applications in the industry, such as the in situ decontamination of xenobiotics.
Assuntos
Amiloide/química , Hidrolases/química , Amiloide/síntese química , Amiloide/metabolismo , Animais , Domínio Catalítico , Humanos , Hidrolases/síntese química , Hidrolases/metabolismoRESUMO
Giardiasis represents a latent problem in public health due to the exceptionally pathogenic strategies of the parasite Giardia lamblia for evading the human immune system. Strains resistant to first-line drugs are also a challenge. Therefore, new antigiardial therapies are urgently needed. Here, we tested giardial arginine deiminase (GlADI) as a target against giardiasis. GlADI belongs to an essential pathway in Giardia for the synthesis of ATP, which is absent in humans. In silico docking with six thiol-reactive compounds was performed; four of which are approved drugs for humans. Recombinant GlADI was used in enzyme inhibition assays, and computational in silico predictions and spectroscopic studies were applied to follow the enzyme's structural disturbance and identify possible effective drugs. Inhibition by modification of cysteines was corroborated using Ellman's method. The efficacy of these drugs on parasite viability was assayed on Giardia trophozoites, along with the inhibition of the endogenous GlADI. The most potent drug against GlADI was assayed on Giardia encystment. The tested drugs inhibited the recombinant GlADI by modifying its cysteines and, potentially, by altering its 3D structure. Only rabeprazole and omeprazole decreased trophozoite survival by inhibiting endogenous GlADI, while rabeprazole also decreased the Giardia encystment rate. These findings demonstrate the potential of GlADI as a target against giardiasis.
Assuntos
Giardia lamblia/efeitos dos fármacos , Giardíase/tratamento farmacológico , Hidrolases/metabolismo , Animais , Antiprotozoários/farmacologia , Simulação por Computador , Cisteína/química , Avaliação Pré-Clínica de Medicamentos/métodos , Reposicionamento de Medicamentos/métodos , Giardia lamblia/patogenicidade , Giardíase/imunologia , Tiomalato Sódico de Ouro/farmacologia , Humanos , Hidrolases/efeitos dos fármacos , Hidrolases/ultraestrutura , Omeprazol/farmacologia , Inibidores da Bomba de Prótons/farmacologia , Rabeprazol , Tiamina/análogos & derivados , Tiamina/farmacologia , Trofozoítos/efeitos dos fármacosRESUMO
OBJECTIVES: This study was aimed at assessing the anti-arthritic effects of hesperidin on the inflammatory markers in serum/plasma, ectoenzymes activity in platelet, reactive oxygen species (ROS), apoptosis and cell cycle in bone marrow cells of a rat model of arthritis. METHODS: Fifty-six adult female Wistar rats (245-274 g) were grouped into eight of seven rats each: control rats given normal saline or 40 mg/kg of hesperidin or 80 mg/kg of hesperidin, 0.2 mg/kg of dexamethasone, arthritic rats given normal saline, or 40 mg/kg of hesperidin or 80 mg/kg of hesperidin, and 0.2 mg/kg of dexamethasone. Myeloperoxidase and nitrate plus nitrite levels were evaluated in the plasma and serum, respectively. The ecto-nucleoside triphosphate diphosphohydrolases, ecto-5'-nucleotidase and ecto-adenosine deaminase activities were assessed in platelets. Subsequently, the cells of the bone marrow were obtained, and the assays for ROS, apoptosis and cell cycle were evaluated using flow cytometry. KEY FINDINGS: The results showed that hesperidin mitigated inflammation, modulated adenosine nucleotides and nucleoside hydrolysing enzymes and levels, minimized ROS intracellularly, attenuated apoptotic process and activated cell cycle arrest in arthritic rat. CONCLUSION: This study suggests that hesperidin could be a natural and promising anti-inflammatory compound for the management of arthritis.
Assuntos
Anti-Inflamatórios/farmacologia , Artrite Experimental , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Citrus/química , Hesperidina/farmacologia , Hidrolases/metabolismo , Inflamação , 5'-Nucleotidase/metabolismo , Adenosina Desaminase/metabolismo , Animais , Anti-Inflamatórios/uso terapêutico , Apoptose , Artrite Experimental/tratamento farmacológico , Artrite Experimental/metabolismo , Artrite Experimental/patologia , Artrite Reumatoide/tratamento farmacológico , Artrite Reumatoide/metabolismo , Artrite Reumatoide/patologia , Feminino , Adjuvante de Freund , Hesperidina/uso terapêutico , Inflamação/metabolismo , Inflamação/prevenção & controle , Proteínas de Membrana/metabolismo , Nucleosídeo-Trifosfatase/metabolismo , Fitoterapia , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Pirofosfatases , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismoRESUMO
Recently, a bacterium strain of Ideonella sakaiensis was identified with the uncommon ability to degrade the poly(ethylene terephthalate) (PET). The PETase from I. sakaiensis strain 201-F6 (IsPETase) catalyzes the hydrolysis of PET converting it to mono(2-hydroxyethyl) terephthalic acid (MHET), bis(2-hydroxyethyl)-TPA (BHET), and terephthalic acid (TPA). Despite the potential of this enzyme for mitigation or elimination of environmental contaminants, one of the limitations of the use of IsPETase for PET degradation is the fact that it acts only at moderate temperature due to its low thermal stability. Besides, molecular details of the main interactions of PET in the active site of IsPETase remain unclear. Herein, molecular docking and molecular dynamics (MD) simulations were applied to analyze structural changes of IsPETase induced by PET binding. Results from the essential dynamics revealed that the ß1-ß2 connecting loop is very flexible. This loop is located far from the active site of IsPETase and we suggest that it can be considered for mutagenesis to increase the thermal stability of IsPETase. The free energy landscape (FEL) demonstrates that the main change in the transition between the unbound to the bound state is associated with the ß7-α5 connecting loop, where the catalytic residue Asp206 is located. Overall, the present study provides insights into the molecular binding mechanism of PET into the IsPETase structure and a computational strategy for mapping flexible regions of this enzyme, which can be useful for the engineering of more efficient enzymes for recycling plastic polymers using biological systems.
Assuntos
Proteínas de Bactérias/metabolismo , Burkholderiales/metabolismo , Hidrolases/metabolismo , Polietilenotereftalatos/metabolismo , Biocatálise , HidróliseRESUMO
Two trials were conducted to evaluate the effects of soyabean meal replacement by maize distillers dried grains with solubles (DDGS) in diets for pacu juveniles. Five diets were formulated with 0, 100, 200, 300 and 400 g of DDGS/kg diet replacing up to total dietary soyabean meal. In trial 1, the experimental diets were fed to five groups of fish to evaluate the apparent digestibility coefficients (ADC). In trial 2, four groups of fish were fed each experimental diet for 100 d to evaluate the effects of these diets on digestive enzyme activity, intestine oxidative stress and intestine morphology. The ADC of DM and energy was reduced with dietary DDGS inclusion, while the ADC of lipids was increased, and no differences were observed for the ADC of protein. Independent of dietary treatment, pH increased from anterior to the distal intestine with dietary DDGS inclusion. Digestive enzyme activities were higher on anterior than the distal intestine. Dietary DDGS decreased lipase, amylase, chymotrypsin and trypsin activities, while no differences were observed for total protease activity. Intestine glucose-6-phosphate dehydrogenase was reduced in fish fed the DDGS diets, while catalase activity increased. Lipid peroxidation was lower in fish fed DDGS diets than the control. Intestine histomorphology improved with dietary DDGS inclusion. Overall, the negative effects of soyabean meal could be decreased by dietary replacement with maize DDGS which may have a prebiotic effect, improving intestine health.
Assuntos
Ração Animal , Dieta , Digestão , Grão Comestível , Peixes/fisiologia , Intestinos/anatomia & histologia , Intestinos/fisiologia , Fenômenos Fisiológicos da Nutrição Animal , Animais , Gorduras na Dieta/metabolismo , Peixes/anatomia & histologia , Concentração de Íons de Hidrogênio , Hidrolases/metabolismo , Mucosa Intestinal/anatomia & histologia , Intestinos/enzimologia , Metabolismo dos Lipídeos , Peroxidação de Lipídeos , Oxirredutases/metabolismo , Peptídeo Hidrolases/metabolismo , Glycine max , Zea maysRESUMO
BACKGROUND: Quizalofop-p-ethyl (QPE), a unitary R configuration aromatic oxyphenoxypropionic acid ester (AOPP) herbicide, was widely used and had led to detrimental environmental effects. For finding the QPEdegrading bacteria and promoting the biodegradation of QPE, a series of studies were carried out. RESULTS: A QPE-degrading bacterial strain YC-XJ1 was isolated from desert soil and identified as Methylobacterium populi, which could degrade QPE with methanol by cometabolism. Ninety-seven percent of QPE (50 mg/L) could be degraded within 72 h under optimum biodegradation condition of 35°C and pH 8.0. The maximum degradation rate of QPE was 1.4 mg/L/h, and the strain YC-XJ1 exhibited some certain salinity tolerance. Two novel metabolites, 2-hydroxy-6-chloroquinoxaline and quinoxaline, were found by high-performance liquid chromatography/mass spectroscopy analysis. The metabolic pathway of QPE was predicted. The catalytic efficiency of strain YC-XJ1 toward different AOPPs herbicides in descending order was as follows: haloxyfop-pmethyl ≈ diclofop-methyl ≈ fluazifop-p-butyl N clodinafop-propargyl N cyhalofop-butyl N quizalofop-p-ethyl N fenoxaprop-p-ethyl N propaquizafop N quizalofop-p-tefuryl. The genome of strain YC-XJ1 was sequenced using a combination of PacBio RS II and Illumina platforms. According to the annotation result, one α/ß hydrolase gene was selected and named qpeh1, for which QPE-degrading function has obtained validation. Based on the phylogenetic analysis and multiple sequence alignment with other QPE-degrading esterases reported previously, the QPEH1 was clustered with esterase family V. CONCLUSION: M. populi YC-XJ1 could degrade QPE with a novel pathway, and the qpeh1 gene was identified as one of QPE-degrading esterase gene.