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1.
Chem Biol Interact ; 313: 108821, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31525342

RESUMO

Decrease in the bioavailability of vasoactive nitric oxide (NO), derived from the endothelial nitric oxide synthase (NOS3), underlines vascular endothelial damage. Our expanding knowledge on mature red blood cells (RBCs) makes it supposable that RBCs might contribute to vascular function and integrity via their active NO synthetizing system (RBC-NOS3). This "rescue" mechanism of RBCs could be especially important during pregnancy with smoking habit, when smoking acts as an additional stressor and causes active change in the redox status. In this study RBC populations of 82 non-smoking (RBC-NS) and 75 smoking (RBC-S) pregnant women were examined. Morphological variants were followed by confocal microscopy and quantified by a microscopy based intelligent analysis software. Fluorescence activated cell sorting was used to examine the translational and posttranslational regulation of RBC-NOS, Arginase-1 and the formation of the major product of lipid peroxidation, 4-hydroxy-2-nonenal. To survey the rheological parameters of RBCs like elasticity and plasticity atomic force microscopy-based measurement was applied. Significant morphological and functional differences of RBCs were found between the non-smoking and smoking groups. The phenotypic variations in RBC-S population, even the characteristic biconcave disc-shaped cells, could be connected to impaired NOS3 activation and are compromised in their physiological properties. Membrane lipid studies reveal an elevated lipid oxidation state well paralleled with the changed elastic and plastic activities. These features can form a basic tool in the prenatal health screening conditions; hence the compensatory mechanism of RBC-S population completely fails to sense and rescue the acute oxidative stress conditions.


Assuntos
Arginase/metabolismo , Eritrócitos/metabolismo , Óxido Nítrico Sintase Tipo III/metabolismo , Óxido Nítrico/metabolismo , Fumar/efeitos adversos , Adulto , Aldeídos/metabolismo , Estudos de Casos e Controles , Morte Celular/efeitos dos fármacos , Eritrócitos/efeitos dos fármacos , Feminino , Humanos , Peroxidação de Lipídeos/efeitos dos fármacos , Microscopia de Força Atômica , Gravidez
2.
Cancer Sci ; 110(9): 2856-2866, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31314163

RESUMO

4-Hydroxynonenal (HNE) is an important product of plasma membrane lipid peroxidation, which is a cause of cell and tissue injury. Mitochondrial DNA (mtDNA)-depleted ρ0 cells were established using human cervical cancer and oral squamous cell carcinoma cell lines. We investigated the effect of reactive oxygen species in ρ0 cells, especially the mechanism of hydrogen peroxide (H2 O2 )-mediated cell death. These cell were subjected to high oxidative stress and, compared with their parental cells, showed greater sensitivity to H2 O2 and high lipid peroxidation. Upregulation of HNE in the plasma membrane was observed prior to the increase in intracellular H2 O2 . The amount of oxidized lipid present changed H2 O2 permeability and administration of oxidized lipid led to further cell death after treatment with H2 O2 . Expression levels of lipoxygenase ALOX genes (ie ALOX5, ALOX12, and ALOX15) were upregulated in ρ0 cells, as were expression levels of ALOX12 and ALOX15 proteins. ALOX5 protein was mainly distributed in the nucleus, while ALOX12 and ALOX15 proteins were distributed in the nucleus and the cytoplasm. Although expression of COX2 gene was upregulated, its protein expression did not increase. ALOX (especially ALOX15) may be involved in the sensitivity of cancer cells to treatment. These data offer promise for the development of novel anticancer agents by altering the oxidation state of the plasma membrane. Our results showed that lipid peroxidation status is important for H2 O2 sensitivity and that ALOX15 is involved in lipid peroxidation status.


Assuntos
Apoptose/efeitos dos fármacos , Permeabilidade da Membrana Celular/genética , DNA Mitocondrial/genética , Peróxido de Hidrogênio/administração & dosagem , Peroxidação de Lipídeos/genética , Neoplasias/patologia , Aldeídos/metabolismo , Araquidonato 15-Lipoxigenase/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Resistencia a Medicamentos Antineoplásicos , Complexo de Proteínas da Cadeia de Transporte de Elétrons/genética , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Peróxido de Hidrogênio/farmacocinética , Mitocôndrias/genética , Mitocôndrias/metabolismo , Neoplasias/tratamento farmacológico , Neoplasias/genética , Estresse Oxidativo/efeitos dos fármacos , Éteres Fosfolipídicos/administração & dosagem , Regulação para Cima
3.
Chem Commun (Camb) ; 55(62): 9080-9083, 2019 Aug 11.
Artigo em Inglês | MEDLINE | ID: mdl-31287110

RESUMO

New strategies are required for the discovery of unknown bioactive molecules produced by gut microbiota in the human host. Herein, we utilize a chemoselective probe immobilized to magnetic beads for analysis of carbonyls in human fecal samples. We identified 112 metabolites due to femtomole analysis and an increased mass spectrometric sensitivity by up to six orders of magnitude.


Assuntos
Aldeídos/análise , Fezes/química , Microbioma Gastrointestinal/fisiologia , Cetonas/análise , Sondas Moleculares/análise , Sondas Moleculares/química , Aldeídos/metabolismo , Fezes/microbiologia , Feminino , Humanos , Cetonas/metabolismo , Masculino , Sondas Moleculares/síntese química , Estrutura Molecular , Tamanho da Partícula , Propriedades de Superfície
4.
Chem Commun (Camb) ; 55(52): 7498-7501, 2019 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-31187106

RESUMO

We explored a collection of 2-deoxyribose-5-phosphate aldolases (DERAs) from biodiversity for their nucleophile substrate promiscuity. The DERAs were screened using as nucleophiles propanone, propanal, cyclobutanone, cyclopentanone, dihydroxyacetone, and glycolaldehyde with l-glyceraldehyde-3-phosphate as an electrophile in aldol addition. A DERA from Arthrobacter chlorophenolicus (DERAArthro) efficiently allowed the synthesis of the corresponding aldol adducts in good yields, displaying complementarity in terms of configuration and substrate specificity with fructose-6-phosphate aldolase, the only previously known aldolase with a large nucleophile tolerance.


Assuntos
Aldeído Liases/metabolismo , Proteínas de Bactérias/metabolismo , Aldeído Liases/genética , Aldeídos/química , Aldeídos/metabolismo , Arthrobacter/enzimologia , Proteínas de Bactérias/genética , Biocatálise , Biodiversidade , Escherichia coli/enzimologia , Gliceraldeído 3-Fosfato/metabolismo , Especificidade por Substrato
5.
Food Chem Toxicol ; 131: 110529, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31150784

RESUMO

The health promoting effects of extra virgin olive oil (EVOO) relate to its unique repertoire of phenolic compounds. Here, we used a chemoinformatics approach to computationally identify endogenous ligands and assign putative biomolecular targets to oleacein, one of the most abundant secoiridoids in EVOO. Using a structure-based virtual profiling software tool and reference databases containing more than 9000 binding sites protein cavities, we identified 996 putative oleacein targets involving more than 700 proteins. We subsequently identified the high-level functions of oleacein in terms of biomolecular interactions, signaling pathways, and protein-protein interaction (PPI) networks. Delineation of the oleacein target landscape revealed that the most significant modules affected by oleacein were associated with metabolic processes (e.g., glucose and lipid metabolism) and chromatin-modifying enzymatic activities (i.e., histone post-translational modifications). We experimentally confirmed that, in a low-micromolar physiological range (<20 µmol/l), oleacein was capable of inhibiting the catalytic activities of predicted metabolic and epigenetic targets including nicotinamide N-methyltransferase, ATP-citrate lyase, lysine-specific demethylase 6A, and N-methyltransferase 4. Our computational de-orphanization of oleacein provides new mechanisms through which EVOO biophenols might operate as chemical prototypes capable of modulating the biologic machinery of healthy aging.


Assuntos
Aldeídos/metabolismo , Fenóis/metabolismo , Proteômica/métodos , ATP Citrato (pro-S)-Liase/química , ATP Citrato (pro-S)-Liase/metabolismo , Aldeídos/química , Domínio Catalítico , Ensaios Enzimáticos , Epigenômica/métodos , Ontologia Genética/estatística & dados numéricos , Histona Desmetilases/química , Histona Desmetilases/metabolismo , Humanos , Informática/métodos , Metiltransferases/química , Metiltransferases/metabolismo , Simulação de Acoplamento Molecular , Nicotinamida N-Metiltransferase/química , Nicotinamida N-Metiltransferase/metabolismo , Olea/química , Azeite de Oliva/química , Fenóis/química , Ligação Proteica , Mapeamento de Interação de Proteínas , Software
6.
Environ Pollut ; 246: 904-913, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31159140

RESUMO

Tris(1,3-dichloro-2-propyl) phosphate (TDCPP) has been frequently detected in environmental media and biological samples. However, knowledge of its adverse health consequences is limited. In the current study, Caenorhabditis elegans (C. elegans, L1 larvae) were exposed to TDCPP at environmentally relevant concentrations (control, 0.1, 1, 100 and 1000 µg L-1) for 72 h to explore any association between TDCPP and the aging process. Some of the degenerative age-related indicators were observed, including locomotion behaviors and lifespan. As crucial biomarkers of aging, the accumulation of lipofuscin, and lipid peroxidation (LPO) products exemplified by 4-hydroxynon-2-enal (4-HNE) were detected. This product forms as a result of oxidative stress, as confirmed by an N-acetyl-L-cysteine (NAC) pharmacological assay. Moreover, a significant increase in reactive oxide species (ROS) production in a dose-dependent manner using a fluorescent probe was observed. For the underlying molecular mechanism of the above aging phenotypes, significantly upregulated transcription of genes related to antioxidant systems, especially a subset of glutathione S-transferase (gst-5, gst-6, gst-9, gst-10, gst-19, gst-24, gst-26, gst-29, gst-33, and gst-38), was found by RNA-Seq and further confirmed by RT-qPCR. The elevated glutathione S-transferase (GST) was attributed to the significant increase in 4-HNE because mutations in gst-5 and gst-24 inhibited the conjugation of GSTs with 4-HNE. Therefore, GST play an indispensable role in the detoxification process of TDCPP exposure and further confirmed LPO accumulation at the molecular mechanism level. In conclusion, TDCPP accelerated the aging process induced by the LPO products, 4-HNE, response to reactive oxidative species in C. elegans.


Assuntos
Envelhecimento/efeitos dos fármacos , Aldeídos/metabolismo , Poluentes Ambientais/toxicidade , Compostos Organofosforados/toxicidade , Espécies Reativas de Oxigênio/metabolismo , Envelhecimento/genética , Envelhecimento/metabolismo , Animais , Caenorhabditis elegans/fisiologia , Glutationa Transferase/genética , Glutationa Transferase/metabolismo , Peroxidação de Lipídeos , Estresse Oxidativo
7.
Food Chem ; 293: 120-126, 2019 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-31151591

RESUMO

The effects of the natural ingredients Angelica sinensis (AS) and Codonopsis pilosula (CP) on the shelf life of chicken seasoning were investigated. Color differences and sensory evaluation were used to indicate sensory differences. Changes in volatiles were monitored. The rate of increase in the color value a* of the AS and CP samples was lower than that in the control. Rancid flavor appeared later in the AS and CP samples than in the control. The levels of aldehydes, ketones, and alkenes increased during storage. A kinetic model was built based on the proportion of aldehydes (main marker), to predict shelf life. The predicted shelf life at room temperature was 60 days for the control, 114 days for AS, and 89 days for CP. The shelf life of chicken seasoning could be prolonged with AS and CP. This kinetic model can be used to predict the shelf life of chicken seasoning.


Assuntos
Aromatizantes/química , Aldeídos/química , Aldeídos/metabolismo , Alcenos/química , Alcenos/metabolismo , Angelica sinensis/química , Angelica sinensis/metabolismo , Animais , Galinhas , Codonopsis/química , Codonopsis/metabolismo , Cor , Aromatizantes/metabolismo , Armazenamento de Alimentos , Cetonas/química , Cetonas/metabolismo , Cinética , Compostos Orgânicos Voláteis/química , Compostos Orgânicos Voláteis/metabolismo
8.
Biochemistry (Mosc) ; 84(Suppl 1): S206-S224, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31213203

RESUMO

Reactive carbonyl compounds (RCC) are a group of compounds with clearly pronounced electrophilic properties that facilitate their spontaneous reactions with numerous nucleophilic reaction sites in proteins, lipids, and nucleic acids. The biological functions of RCC are determined by their concentration and governed by the hormesis (biphasic reaction) principle. At low concentrations, RCC act as signaling molecules activating defense systems against xenobiotics and oxidizers, and at high concentrations, they exhibit the cytotoxic effect. RCC participate in the formation of cell adaptive response via intracellular signaling pathways involving regulation of gene expression and cytoplasmic mechanisms related to the structure-functional rearrangements of proteins. Special attention in this review is given to the functioning of electrophiles as mediators of cell general adaption syndrome manifested as the biphasic response. The hypothesis is proposed that electrophilic signaling can be a proto-signaling system.


Assuntos
Aldeídos/metabolismo , Radicais Livres/metabolismo , Cetonas/metabolismo , Mutagênese/fisiologia , Estresse Oxidativo/fisiologia , Transdução de Sinais/fisiologia , Animais , Humanos , Oxirredução , Processamento de Proteína Pós-Traducional
9.
Chemistry ; 25(43): 10156-10164, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31136019

RESUMO

A new protocol based on lipase-catalyzed tandem reaction toward α,ß-enones/enoesters is presented. For the synthesis of the desired products the tandem process based on enzyme-catalyzed hydrolysis and Knoevenagel reaction starting from enol acetates and aldehyde is developed. The relevant impact of the reaction conditions including organic solvent, enzyme type, and temperature on the course of the reaction was revealed. It was shown that controllable release of the active methylene compound from the corresponding enol carboxylate ensured by enzymatic reaction diminishes significantly the formation of the unwanted co-products. Furthermore, this protocol was extended by including a second tandem chemoenzymatic transformation engaging various aldehyde precursors. After a careful optimization of the reaction conditions, the target products were obtained with yields up to 86 % and with excellent E/Z-selectivity.


Assuntos
Aldeídos/metabolismo , Lipase/metabolismo , Aldeídos/química , Animais , Biocatálise , Hidrólise , Lipase/química , Pancrelipase/metabolismo , Pentanonas/química , Pentanonas/metabolismo , Estereoisomerismo , Suínos
10.
BMC Plant Biol ; 19(1): 187, 2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31064322

RESUMO

BACKGROUND: It is important to explore renewable alternatives (e.g. biofuels) that can produce energy sources to help reduce reliance on fossil oils, and reduce greenhouse gases and waste solids resulted from fossil oils consumption. Camelina sativa is an oilseed crop which has received increasing attention due to its short life cycle, broader adaptation regions, high oil content, high level of omega-3 unsaturated fatty acids, and low-input requirements in agriculture practices. To expand its Camelina production areas into arid regions, there is a need to breed for new drought-tolerant cultivars. Leaf cuticular wax is known to facilitate plant development and growth under water-limited conditions. Dissecting the genetic loci underlying leaf cuticular waxes is important to breed for cultivars with improved drought tolerance. RESULTS: Here we combined phenotypic data and single nucleotide polymorphism (SNP) data from a spring C. sativa diversity panel using genotyping-by-sequencing (GBS) technology, to perform a large-scale genome-wide association study (GWAS) on leaf wax compositions. A total of 42 SNP markers were significantly associated with 15 leaf wax traits including major wax components such as total primary alcohols, total alkanes, and total wax esters as well as their constituents. The vast majority of significant SNPs were associated with long-chain carbon monomers (carbon chain length longer than C28), indicating the important effects of long-chain carbon monomers on leaf total wax biosynthesis. These SNP markers are located on genes directly or indirectly related to wax biosynthesis such as maintaining endoplasmic reticulum (ER) morphology and enabling normal wax secretion from ER to plasma membrane or Golgi network-mediated transport. CONCLUSIONS: These loci could potentially serve as candidates for the genetic control involved in intracellular wax transport that might directly or indirectly facilitate leaf wax accumulation in C. sativa and can be used in future marker-assisted selection (MAS) to breed for the cultivars with high wax content to improve drought tolerance.


Assuntos
Brassicaceae/genética , Folhas de Planta/química , Polimorfismo de Nucleotídeo Único , Ceras/química , Ceras/metabolismo , Álcoois/metabolismo , Aldeídos/metabolismo , Algoritmos , Alcanos/metabolismo , Transporte Biológico/genética , Genética Populacional , Estudo de Associação Genômica Ampla , Desequilíbrio de Ligação , Fenótipo , Folhas de Planta/genética
11.
Chem Biol Interact ; 306: 104-109, 2019 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-30998906

RESUMO

Autophagy is a dynamic recycling process that eliminates damaged proteins and cellular organelles to maintain cellular homeostasis. Aldose reductase (AR) catalyzes conversion of glucose to sorbitol. It also catalyzes the reduction of a broad array of saturated and unsaturated aldehydes. Recently we demonstrated that deletion of AR promotes pathological cardiac remodeling via excessive autophagy; however, the role of AR in starvation-induced autophagy has not been determined. To determine the role of AR in starvation-induced autophagy, WTC57/Bl6 mice were pretreated with the AR inhibitor sorbinil (0.2 g/L for 48 h) in drinking water, followed by 24 h fasting. We found that the sorbinil pretreatment in fed mice did not affect blood glucose levels, whereas, it decreased the blood glucose levels in fasting mice. In comparison with fed mice, the LC3II formation and LCII/LCI ratio were increased in the fasted mice hearts and sorbinil pretreatment further enhanced LC3II formation and LC3II/LC3I ratios in these hearts. Fasting-induced autophagy coincided with AMPK activation in the sorbinil pretreated fasted mice hearts. Autophagy and activation of AMPK was also induced in the gastrocnemius skeletal muscle of sorbinil pre-treated fasted mice. Induction of autophagy in the cardiac tissues of sorbinil pretreated fasted mice was accompanied by increased clearance of 4-hydroxytrans-2-nonenal-protein adducts. Taken together, these results indicate that the inhibition of AR during fasting activates autophagic response, increases clearance of aldehyde-protein adducts, which could serve as a mechanism to maintain cellular homeostasis during starvation.


Assuntos
Aldeído Redutase/antagonistas & inibidores , Aldeídos/metabolismo , Autofagia/efeitos dos fármacos , Imidazolidinas/farmacologia , Inanição , Aldeído Redutase/metabolismo , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL
12.
Chem Biol Interact ; 305: 86-97, 2019 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-30928398

RESUMO

Substrate inhibition by the aldehyde has been observed for decades in NAD(P)+-dependent aldehyde dehydrogenase (ALDH) enzymes, which follow a Bi Bi ordered steady-state kinetic mechanism. In this work, by using theoretical simulations of different possible substrate inhibition mechanisms in monosubstrate and Bi Bi ordered steady-state reactions, we explored the kind and extent of errors arising when estimating the kinetic parameters and determining the kinetic mechanisms if substrate inhibition is intentionally or unintentionally ignored. We found that, in every mechanism, fitting the initial velocity data of apparently non-inhibitory substrate concentrations to a rectangular hyperbola produces important errors, not only in the estimation of Vmax values, which were underestimated as expected, but, surprisingly, even more in the estimation of Km values, which led to overestimation of the Vmax/Km values. We show that the greater errors in Km arises from fitting data that do experience substrate inhibition, although it may not be evident, to a Michaelis-Menten equation, which causes overestimation of the data at low substrate concentrations. Similarly, we show that if substrate inhibition is not fully assessed when inhibitors are evaluated, the estimated inhibition constants will have significant errors, and the type of inhibition could be grossly mistaken. We exemplify these errors with experimental results obtained with the betaine aldehyde dehydrogenase from spinach showing the errors predicted by the theoretical simulations and that these errors are increased in the presence of NADH, which in this enzyme favors aldehyde substrate inhibition. Therefore, we strongly recommend assessing substrate inhibition by the aldehyde in every ALDH kinetic study, particularly when inhibitors are evaluated. The common practices of using an apparently non-inhibitory concentration range of the aldehyde or a single high concentration of the aldehyde or the coenzyme when varying the other to determine true kinetic parameters should be abandoned.


Assuntos
Aldeído Desidrogenase/metabolismo , Aldeídos/metabolismo , Proteínas de Plantas/metabolismo , Proteínas Recombinantes/metabolismo , Aldeído Desidrogenase/antagonistas & inibidores , Aldeído Desidrogenase/genética , Aldeídos/química , Cinética , NAD/metabolismo , Proteínas de Plantas/antagonistas & inibidores , Proteínas de Plantas/genética , Proteínas Recombinantes/genética , Spinacia oleracea/enzimologia , Especificidade por Substrato
13.
J Sci Food Agric ; 99(11): 5028-5034, 2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-30989657

RESUMO

BACKGROUND: The influence of heat-induced structural modifications of grass carp myofibrillar protein (MP) on its ability to bind to selected aldehydes (hexanal, heptanal, octanal and nonanal) was investigated. The interactions of MP and flavor compounds were investigated using HS-GC-MS, intrinsic fluorescence spectra, Raman spectra, SDS-PAGE, turbidity, total sulfhydryl content and surface hydrophobicity. RESULTS: The ability to bind to aldehydes was strongly influenced by changes in the structure and surface of proteins during the heating process (0-30 min). During the first 0-10 min of heating, the flavor-binding ability increased, which is likely attributable to increased surface hydrophobicity and total sulfhydryl content, and to the unfolding of secondary structures of MP by exposure to reactive amino acids, sulfhydryl groups and hydrophobic bonding sites. Nevertheless, lengthy heating (>10 min) caused protein refolding and accelerated aggregation of protein, thus reducing hydrophobic interactions and weakening the resultant capacity of MP to bind to flavor compounds. CONCLUSION: The results suggested that hydrophobic interactions were enhanced upon short-term heating, whereas long-term heating weakend them. The results provide information concerning improvement of the flavor profile of freshwater fish surimi products. © 2019 Society of Chemical Industry.


Assuntos
Aldeídos/metabolismo , Cyprinidae , Temperatura Alta , Proteínas Musculares/metabolismo , Miofibrilas/química , Animais , Eletroforese em Gel de Poliacrilamida , Cromatografia Gasosa-Espectrometria de Massas , Interações Hidrofóbicas e Hidrofílicas , Estrutura Molecular , Proteínas Musculares/química , Ligação Proteica , Espectrometria de Fluorescência , Análise Espectral Raman , Compostos de Sulfidrila/análise , Paladar
14.
Appl Microbiol Biotechnol ; 103(9): 3807-3817, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30877354

RESUMO

Enzyme-mediated synthesis of pharmaceutical compounds is a 'green' alternative to traditional synthetic chemistry, and microbial engineering opens up the possibility of using whole cells as mini-factories. Whole-cell biocatalysis reduces cost by eliminating expensive enzyme purification and cofactor addition steps, as well as resulting in increased enzyme stability. Haloferax volcanii is a model halophilic archaeon encoding highly salt and organic solvent tolerant enzymes such as alcohol dehydrogenase (HvADH2), which catalyses the reduction of aldehydes and ketone in the presence of NADPH/NADH cofactor. A H. volcanii strain for constitutive HvADH2 expression was generated using a strong synthetic promoter (p.syn). The strain was immobilised in calcium alginate beads and repeatedly used as a whole-cell biocatalyst. The reduction of acetophenone, used as test substrate, was very successful and high yields were detected from immobilised whole cells over repeated biotransformation cycles. The immobilised H. volcanii retained stability and high product yields after 1 month of storage at room temperature. This newly developed system offers halophilic enzyme expression in its native environment, high product yield, stability and reusability without the addition of any expensive NADPH/NADH cofactor. This is the first report of whole cell-mediated biocatalysis by the halophilic archaeon H. volcanii.


Assuntos
Álcool Desidrogenase/química , Proteínas Arqueais/química , Haloferax volcanii/metabolismo , Sais/metabolismo , Acetofenonas/metabolismo , Álcool Desidrogenase/genética , Álcool Desidrogenase/metabolismo , Aldeídos/metabolismo , Proteínas Arqueais/genética , Proteínas Arqueais/metabolismo , Biocatálise , Células Imobilizadas/química , Células Imobilizadas/enzimologia , Células Imobilizadas/metabolismo , Estabilidade Enzimática , Expressão Gênica , Haloferax volcanii/química , Haloferax volcanii/enzimologia , Cetonas/metabolismo , NADP/metabolismo
15.
Planta ; 249(6): 1823-1836, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30847571

RESUMO

MAIN CONCLUSION: The enzymes HaKCS1 and HaKCS2 are expressed in sunflower seeds and contribute to elongation of C18 fatty acids, resulting in the C20-C24 fatty acids in sunflower oil. Most plant fatty acids are produced by plastidial soluble fatty acid synthases that produce fatty acids of up to 18 carbon atoms. However, further acyl chain elongations can take place in the endoplasmic reticulum, catalysed by membrane-bound synthases that act on acyl-CoAs. The condensing enzymes of these complexes are the ketoacyl-CoA synthase (KCSs), responsible for the synthesis of very long chain fatty acids (VLCFAs) and their derivatives in plants, these including waxes and cuticle hydrocarbons, as well as fatty aldehydes. Sunflower seeds accumulate oil that contains around 2-3% of VLCFAs and studies of the fatty acid elongase activity in developing sunflower embryos indicate that two different KCS isoforms drive the synthesis of these fatty acids. Here, two cDNAs encoding distinct KCSs were amplified from RNAs extracted from developing sunflower embryos and named HaKCS1 and HaKCS2. These genes are expressed in developing seeds during the period of oil accumulation and they are clear candidates to condition sunflower oil synthesis. These two KCS cDNAs complement a yeast elongase null mutant and when expressed in yeast, they alter the host's fatty acid profile, proving the encoded KCSs are functional. The structure of these enzymes was modelled and their contribution to the presence of VLCFAs in sunflower oil is discussed based on the results obtained.


Assuntos
Acetiltransferases/metabolismo , Helianthus/enzimologia , Modelos Estruturais , Óleo de Girassol/metabolismo , Acetiltransferases/química , Acetiltransferases/genética , Acil Coenzima A/metabolismo , Aldeídos/metabolismo , Sequência de Aminoácidos , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , DNA Complementar/genética , Ácido Graxo Sintases/química , Ácido Graxo Sintases/genética , Ácido Graxo Sintases/metabolismo , Ácidos Graxos/metabolismo , Helianthus/genética , Filogenia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Sementes/enzimologia , Sementes/genética , Alinhamento de Sequência
16.
Oxid Med Cell Longev ; 2019: 2471312, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30906501

RESUMO

Accumulation of oxidative insults on molecular and supramolecular levels could compromise renewal potency and architecture in the aging skin. To examine and compare morphological and ultrastructural changes with redox alterations during chronological skin aging, activities of antioxidant defense (AD) enzymes, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione reductase (GR), thioredoxin reductase (TR), and methionine sulfoxide reductase A (MsrA), and the markers of oxidative damage of biomolecules-4-hydroxynonenal (HNE) and 8-oxoguanine (8-oxoG)-were examined in the rat skin during life (from 3 days to 21 months). As compared to adult 3-month-old skin, higher activities of CAT, GSH-Px, and GR and a decline in expression of MsrA are found in 21-month-old skin. These changes correspond to degenerative changes at structural and ultrastructural levels in epidermal and dermal compartments, low proliferation capacity, and higher levels of HNE-modified protein aldehydes (particularly in basal lamina) and 8-oxoG positivity in nuclei and mitochondria in the sebaceous glands and root sheath. In 3-day-old skin, higher activities of AD enzymes (SOD, CAT, GR, and TR) and MsrA expression correspond to intensive postnatal development and proliferation. In contrast to 21-month-old skin, a high level of HNE in young skin is not accompanied by 8-oxoG positivity or any morphological disturbances. Observed results indicate that increased activity of AD enzymes in elderly rat skin represents the compensatory response to accumulated oxidative damage of DNA and proteins, accompanied by attenuated repair and proliferative capacity, but in young rats the redox changes are necessary and inherent with processes which occur during postnatal skin development. Мorphological and ultrastructurаl changes are in line with the redox profile in the skin of young and old rats.


Assuntos
Envelhecimento/metabolismo , Pele/metabolismo , Pele/patologia , Aldeídos/metabolismo , Animais , Catalase/metabolismo , Proliferação de Células , Colágeno/ultraestrutura , Dano ao DNA , Desoxiguanosina/análogos & derivados , Desoxiguanosina/metabolismo , Fibroblastos/ultraestrutura , Glutationa Peroxidase/metabolismo , Glutationa Redutase/metabolismo , Masculino , Oxirredução , Oxirredutases/metabolismo , Antígeno Nuclear de Célula em Proliferação/metabolismo , Ratos Wistar , Pele/enzimologia , Pele/ultraestrutura , Superóxido Dismutase/metabolismo , Tiorredoxina Dissulfeto Redutase/metabolismo , Fatores de Tempo
17.
Chem Biol Interact ; 305: 21-27, 2019 May 25.
Artigo em Inglês | MEDLINE | ID: mdl-30926318

RESUMO

Voltage-gated potassium (Kv) channels play an essential role in the regulation of membrane excitability and thereby control physiological processes such as cardiac excitability, neural communication, muscle contraction, and hormone secretion. Members of the Kv1 and Kv4 families are known to associate with auxiliary intracellular Kvß subunits, which belong to the aldo-keto reductase superfamily. Electrophysiological studies have shown that these proteins regulate the gating properties of Kv channels. Although the three gene products encoding Kvß proteins are functional enzymes in that they catalyze the nicotinamide adenine dinucleotide phosphate (NAD[P]H)-dependent reduction of a wide range of aldehyde and ketone substrates, the physiological role for these proteins and how each subtype may perform unique roles in coupling membrane excitability with cellular metabolic processes remains unclear. Here, we discuss current knowledge of the enzymatic properties of Kvß proteins from biochemical studies with their described and purported physiological and pathophysiological influences.


Assuntos
Canais de Potássio de Abertura Dependente da Tensão da Membrana/metabolismo , Aldeídos/metabolismo , Animais , Humanos , Sistema Imunitário/metabolismo , Cetonas/metabolismo , Cinética , Miocárdio/metabolismo , NADP/metabolismo , Sistema Nervoso/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana/antagonistas & inibidores
18.
J Agric Food Chem ; 67(14): 3845-3853, 2019 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-30875206

RESUMO

Extra virgin olive oil (EVOO) is the main source of fat in the Mediterranean diet. Phenolic compounds of EVOO, in particular, secoiridoids, are minor components that have generated special interest due to their positive effects on human health, supported by several clinical trials. This review summarizes the most recent findings on the pharmacological properties and action's mechanisms of secoiridoid oleocanthal, focusing attention on inflammation, oxidative stress, cancer, neurodegenerative processes, and rheumatic diseases. Being of relevance to the clinical effects of EVOO intake, the bioavailability and biotransformation of EVOO polyphenols are addressed. Moreover, this review summarizes the factors that may influence the oleocanthal concentration in EVOO. With the growing incidence of age- and lifestyle-related diseases, the current data indicated that the administration of EVOO rich in secoiridoids may be helpful in the prevention or treatment of different pathologies with an inflammatory component. Although promising, the future raises several questions and challenges, which are discussed here. The real beneficial effects of olive oil phenols on human health need to be clarified in new, well-designed clinical studies.


Assuntos
Aldeídos/metabolismo , Inflamação/dietoterapia , Azeite de Oliva/metabolismo , Fenóis/metabolismo , Aldeídos/química , Animais , Estilo de Vida Saudável , Humanos , Inflamação/genética , Inflamação/imunologia , Olea/química , Olea/metabolismo , Azeite de Oliva/química , Fenóis/química
19.
Molecules ; 24(6)2019 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-30889828

RESUMO

The members of the Old Yellow Enzyme (OYE) family are capable of catalyzing the asymmetric reduction of (E/Z)-citral to (R)-citronellal-a key intermediate in the synthesis of L-menthol. The applications of OYE-mediated biotransformation are usually hampered by its insufficient enantioselectivity and low activity. Here, the (R)-enantioselectivity of Old Yellow Enzyme from Saccharomyces cerevisiae CICC1060 (OYE2y) was enhanced through protein engineering. The single mutations of OYE2y revealed that the sites R330 and P76 could act as the enantioselectivity switch of OYE2y. Site-saturation mutagenesis was conducted to generate all possible replacements for the sites R330 and P76, yielding 17 and five variants with improved (R)-enantioselectivity in the (E/Z)-citral reduction, respectively. Among them, the variants R330H and P76C partly reversed the neral derived enantioselectivity from 32.66% e.e. (S) to 71.92% e.e. (R) and 37.50% e.e. (R), respectively. The docking analysis of OYE2y and its variants revealed that the substitutions R330H and P76C enabled neral to bind with a flipped orientation in the active site and thus reverse the enantioselectivity. Remarkably, the double substitutions of R330H/P76M, P76G/R330H, or P76S/R330H further improved (R)-enantioselectivity to >99% e.e. in the reduction of (E)-citral or (E/Z)-citral. The results demonstrated that it was feasible to alter the enantioselectivity of OYEs through engineering key residue distant from active sites, e.g., R330 in OYE2y.


Assuntos
Aldeídos/metabolismo , Engenharia Metabólica/métodos , Monoterpenos/metabolismo , NADPH Desidrogenase/química , Saccharomyces cerevisiae/enzimologia , Sequência de Aminoácidos , Substituição de Aminoácidos , Biocatálise , Modelos Moleculares , Mutagênese/genética , Proteínas Mutantes/metabolismo , NADPH Desidrogenase/metabolismo , Oxirredução , Estereoisomerismo
20.
Chem Biol Interact ; 304: 194-201, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-30768969

RESUMO

Plant cytosolic aldehyde dehydrogenases from family 2 (ALDH2s, EC 1.2.1.3) are non-specific enzymes and participate for example in the metabolism of acetaldehyde or biosynthesis of phenylpropanoids. Plant aminoaldehyde dehydrogenases (AMADHs, ALDH10 family, EC 1.2.1.19) are broadly specific and play an important role in polyamine degradation or production of osmoprotectants. We have tested imidazole and pyrazole carbaldehydes and their alkyl-, allyl-, benzyl-, phenyl-, pyrimidinyl- or thienyl-derivatives as possible substrates of plant ALDH2 and ALDH10 enzymes. Imidazole represents a building block of histidine, histamine as well as certain alkaloids. It also appears in synthetic pharmaceuticals such as imidazole antifungals. Biological compounds containing pyrazole are rare (e.g. pyrazole-1-alanine and pyrazofurin antibiotics) but the ring is often found as a constituent of many synthetic drugs and pesticides. The aim was to evaluate whether aldehyde compounds based on azole heterocycles are oxidized by the enzymes, which would further support their expected role as detoxifying aldehyde scavengers. The analyzed imidazole and pyrazole carbaldehydes were only slowly converted by ALDH10s but well oxidized by cytosolic maize ALDH2 isoforms (particularly by ALDH2C1). In the latter case, the respective Km values were in the range of 10-2000 µmol l-1; the kcat values appeared mostly between 0.1 and 1.0 s-1. The carbaldehyde group at the position 4 of imidazole was oxidized faster than that at the position 2. Such a difference was not observed for pyrazole carbaldehydes. Aldehydes with an aromatic substituent on their heterocyclic ring were oxidized faster than those with an aliphatic substituent. The most efficient of the tested substrates were comparable to benzaldehyde and p-anisaldehyde known as the best aromatic aldehyde substrates of plant cytosolic ALDH2s in vitro.


Assuntos
Aldeído Desidrogenase/metabolismo , Aldeídos/metabolismo , Lycopersicon esculentum/enzimologia , Ervilhas/enzimologia , Zea mays/enzimologia , Aldeídos/química , Imidazóis/química , Imidazóis/metabolismo , Estrutura Molecular , Oxirredução , Pirazóis/química , Pirazóis/metabolismo
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