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1.
Cell Mol Biol Lett ; 29(1): 65, 2024 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-38714951

RESUMO

The engineered clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein (Cas) system is currently widely applied in genetic editing and transcriptional regulation. The catalytically inactivated CasRx (dCasRx) has the ability to selectively focus on the mRNA coding region without disrupting transcription and translation, opening up new avenues for research on RNA modification and protein translation control. This research utilized dCasRx to create a translation-enhancement system for mammals called dCasRx-eIF4GI, which combined eukaryotic translation initiation factor 4G (eIF4GI) to boost translation levels of the target gene by recruiting ribosomes, without affecting mRNA levels, ultimately increasing translation levels of different endogenous proteins. Due to the small size of dCasRx, the dCasRx-eIF4GI translation enhancement system was integrated into a single viral vector, thus optimizing the delivery and transfection efficiency in subsequent applications. Previous studies reported that ferroptosis, mediated by calcium oxalate (CaOx) crystals, significantly promotes stone formation. In order to further validate its developmental potential, it was applied to a kidney stone model in vitro and in vivo. The manipulation of the ferroptosis regulatory gene FTH1 through single-guide RNA (sgRNA) resulted in a notable increase in FTH1 protein levels without affecting its mRNA levels. This ultimately prevented intracellular ferroptosis and protected against cell damage and renal impairment caused by CaOx crystals. Taken together, this study preliminarily validated the effectiveness and application prospects of the dCasRx-eIF4GI translation enhancement system in mammalian cell-based disease models, providing novel insights and a universal tool platform for protein translation research and future therapeutic approaches for nephrolithiasis.


Assuntos
Sistemas CRISPR-Cas , Oxalato de Cálcio , Rim , Animais , Humanos , Masculino , Camundongos , Oxalato de Cálcio/metabolismo , Sistemas CRISPR-Cas/genética , Fator de Iniciação Eucariótico 4G/metabolismo , Fator de Iniciação Eucariótico 4G/genética , Ferritinas , Ferroptose/genética , Edição de Genes/métodos , Células HEK293 , Rim/metabolismo , Rim/patologia , Cálculos Renais/genética , Cálculos Renais/metabolismo , Oxirredutases/metabolismo , Oxirredutases/genética , Biossíntese de Proteínas/genética , RNA Guia de Sistemas CRISPR-Cas/genética , RNA Guia de Sistemas CRISPR-Cas/metabolismo
2.
Appl Microbiol Biotechnol ; 108(1): 323, 2024 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-38713233

RESUMO

Ergot alkaloids (EAs) are a diverse group of indole alkaloids known for their complex structures, significant pharmacological effects, and toxicity to plants. The biosynthesis of these compounds begins with chanoclavine-I aldehyde (CC aldehyde, 2), an important intermediate produced by the enzyme EasDaf or its counterpart FgaDH from chanoclavine-I (CC, 1). However, how CC aldehyde 2 is converted to chanoclavine-I acid (CC acid, 3), first isolated from Ipomoea violacea several decades ago, is still unclear. In this study, we provide in vitro biochemical evidence showing that EasDaf not only converts CC 1 to CC aldehyde 2 but also directly transforms CC 1 into CC acid 3 through two sequential oxidations. Molecular docking and site-directed mutagenesis experiments confirmed the crucial role of two amino acids, Y166 and S153, within the active site, which suggests that Y166 acts as a general base for hydride transfer, while S153 facilitates proton transfer, thereby increasing the acidity of the reaction. KEY POINTS: • EAs possess complicated skeletons and are widely used in several clinical diseases • EasDaf belongs to the short-chain dehydrogenases/reductases (SDRs) and converted CC or CC aldehyde to CC acid • The catalytic mechanism of EasDaf for dehydrogenation was analyzed by molecular docking and site mutations.


Assuntos
Simulação de Acoplamento Molecular , Mutagênese Sítio-Dirigida , Alcaloides de Claviceps/biossíntese , Alcaloides de Claviceps/química , Alcaloides de Claviceps/metabolismo , Aldeídos/metabolismo , Aldeídos/química , Oxirredução , Domínio Catalítico , Oxirredutases/metabolismo , Oxirredutases/genética , Oxirredutases/química
3.
Protein Sci ; 33(6): e4997, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38723110

RESUMO

Rieske oxygenases (ROs) are a diverse metalloenzyme class with growing potential in bioconversion and synthetic applications. We postulated that ROs are nonetheless underutilized because they are unstable. Terephthalate dioxygenase (TPADO PDB ID 7Q05) is a structurally characterized heterohexameric α3ß3 RO that, with its cognate reductase (TPARED), catalyzes the first intracellular step of bacterial polyethylene terephthalate plastic bioconversion. Here, we showed that the heterologously expressed TPADO/TPARED system exhibits only ~300 total turnovers at its optimal pH and temperature. We investigated the thermal stability of the system and the unfolding pathway of TPADO through a combination of biochemical and biophysical approaches. The system's activity is thermally limited by a melting temperature (Tm) of 39.9°C for the monomeric TPARED, while the independent Tm of TPADO is 50.8°C. Differential scanning calorimetry revealed a two-step thermal decomposition pathway for TPADO with Tm values of 47.6 and 58.0°C (ΔH = 210 and 509 kcal mol-1, respectively) for each step. Temperature-dependent small-angle x-ray scattering and dynamic light scattering both detected heat-induced dissociation of TPADO subunits at 53.8°C, followed by higher-temperature loss of tertiary structure that coincided with protein aggregation. The computed enthalpies of dissociation for the monomer interfaces were most congruent with a decomposition pathway initiated by ß-ß interface dissociation, a pattern predicted to be widespread in ROs. As a strategy for enhancing TPADO stability, we propose prioritizing the re-engineering of the ß subunit interfaces, with subsequent targeted improvements of the subunits.


Assuntos
Estabilidade Enzimática , Oxirredutases/química , Oxirredutases/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Modelos Moleculares , Dioxigenases/química , Dioxigenases/metabolismo , Dioxigenases/genética , Temperatura , Escherichia coli/enzimologia , Escherichia coli/genética , Escherichia coli/metabolismo , Polietilenotereftalatos/química , Polietilenotereftalatos/metabolismo , Concentração de Íons de Hidrogênio , Complexo III da Cadeia de Transporte de Elétrons
4.
Planta ; 259(6): 147, 2024 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-38714547

RESUMO

MAIN CONCLUSION: CsNAC086 was found to promote the expression of CsFLS, thus promoting the accumulation of flavonols in Camellia sinensis. Flavonols, the main flavonoids in tea plants, play an important role in the taste and quality of tea. In this study, a NAC TF gene CsNAC086 was isolated from tea plants and confirmed its regulatory role in the expression of flavonol synthase which is a key gene involved in the biosynthesis of flavonols in tea plant. Yeast transcription-activity assays showed that CsNAC086 has self-activation activity. The transcriptional activator domain of CsNAC086 is located in the non-conserved C-terminal region (positions 171-550), while the conserved NAC domain (positions 1-170) does not have self-activation activity. Silencing the CsNAC086 gene using antisense oligonucleotides significantly decreased the expression of CsFLS. As a result, the concentration of flavonols decreased significantly. In overexpressing CsNAC086 tobacco leaves, the expression of NtFLS was significantly increased. Compared with wild-type tobacco, the flavonols concentration increased. Yeast one-hybrid assays showed CsNAC086 did not directly regulate the gene expression of CsFLS. These findings indicate that CsNAC086 plays a role in regulating flavonols biosynthesis in tea plants, which has important implications for selecting and breeding of high-flavonols-concentration containing tea-plant cultivars.


Assuntos
Camellia sinensis , Flavonóis , Regulação da Expressão Gênica de Plantas , Nicotiana , Proteínas de Plantas , Camellia sinensis/genética , Camellia sinensis/metabolismo , Flavonóis/biossíntese , Flavonóis/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Nicotiana/genética , Nicotiana/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Folhas de Planta/metabolismo , Folhas de Planta/genética , Plantas Geneticamente Modificadas
5.
Exp Dermatol ; 33(5): e15101, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38770555

RESUMO

Skin hyperpigmentation is mainly caused by excessive synthesis of melanin; however, there is still no safe and effective therapy for its removal. Here, we found that the dermal freezer was able to improve UVB-induced hyperpigmentation of guinea pigs without causing obvious epidermal damage. We also mimic freezing stimulation at the cellular level by rapid freezing and observed that freezing treatments <2.5 min could not decrease cell viability or induce cell apoptosis in B16F10 and Melan-A cells. Critically, melanin content and tyrosinase activity in two cells were greatly reduced after freezing treatments. The dramatic decrease in tyrosinase activity was associated with the downregulation of MITF, TYR, TRP-1 and TRP-2 protein expression in response to freezing treatments for two cells. Furthermore, our results first demonstrated that freezing treatments significantly reduced the levels of p-GSK3ß and ß-catenin and the nuclear accumulation of ß-catenin in B16F10 and Melan-A cells. Together, these data suggest that fast freezing treatments can inhibit melanogenesis-related gene expression in melanocytes by regulating the Wnt/ß-catenin signalling pathway. The inhibition of melanin production eventually contributed to the improvement in skin hyperpigmentation induced by UVB. Therefore, fast freezing treatments may be a new alternative of skin whitening in the clinic in the future.


Assuntos
Congelamento , Hiperpigmentação , Melaninas , Melanócitos , Monofenol Mono-Oxigenase , Raios Ultravioleta , Via de Sinalização Wnt , beta Catenina , Animais , Melaninas/biossíntese , Melaninas/metabolismo , Melanócitos/metabolismo , Camundongos , Hiperpigmentação/metabolismo , beta Catenina/metabolismo , Monofenol Mono-Oxigenase/metabolismo , Cobaias , Fator de Transcrição Associado à Microftalmia/metabolismo , Sobrevivência Celular , Oxirredutases Intramoleculares/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Apoptose , Oxirredutases/metabolismo , Interferon Tipo I , Proteínas da Gravidez
6.
Protein Sci ; 33(6): e5014, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38747384

RESUMO

A heterodisulfide reductase-like complex (sHdr) and novel lipoate-binding proteins (LbpAs) are central players of a wide-spread pathway of dissimilatory sulfur oxidation. Bioinformatic analysis demonstrate that the cytoplasmic sHdr-LbpA systems are always accompanied by sets of sulfur transferases (DsrE proteins, TusA, and rhodaneses). The exact composition of these sets may vary depending on the organism and sHdr system type. To enable generalizations, we studied model sulfur oxidizers from distant bacterial phyla, that is, Aquificota and Pseudomonadota. DsrE3C of the chemoorganotrophic Alphaproteobacterium Hyphomicrobium denitrificans and DsrE3B from the Gammaproteobacteria Thioalkalivibrio sp. K90mix, an obligate chemolithotroph, and Thiorhodospira sibirica, an obligate photolithotroph, are homotrimers that donate sulfur to TusA. Additionally, the hyphomicrobial rhodanese-like protein Rhd442 exchanges sulfur with both TusA and DsrE3C. The latter is essential for sulfur oxidation in Hm. denitrificans. TusA from Aquifex aeolicus (AqTusA) interacts physiologically with AqDsrE, AqLbpA, and AqsHdr proteins. This is particularly significant as it establishes a direct link between sulfur transferases and the sHdr-LbpA complex that oxidizes sulfane sulfur to sulfite. In vivo, it is unlikely that there is a strict unidirectional transfer between the sulfur-binding enzymes studied. Rather, the sulfur transferases form a network, each with a pool of bound sulfur. Sulfur flux can then be shifted in one direction or the other depending on metabolic requirements. A single pair of sulfur-binding proteins with a preferred transfer direction, such as a DsrE3-type protein towards TusA, may be sufficient to push sulfur into the sink where it is further metabolized or needed.


Assuntos
Proteínas de Bactérias , Oxirredução , Oxirredutases , Enxofre , Sulfurtransferases , Enxofre/metabolismo , Sulfurtransferases/metabolismo , Sulfurtransferases/química , Sulfurtransferases/genética , Oxirredutases/metabolismo , Oxirredutases/química , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética
7.
Nat Commun ; 15(1): 4092, 2024 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-38750010

RESUMO

Nitrous oxide (N2O) is a climate-active gas with emissions predicted to increase due to agricultural intensification. Microbial reduction of N2O to dinitrogen (N2) is the major consumption process but microbial N2O reduction under acidic conditions is considered negligible, albeit strongly acidic soils harbor nosZ genes encoding N2O reductase. Here, we study a co-culture derived from acidic tropical forest soil that reduces N2O at pH 4.5. The co-culture exhibits bimodal growth with a Serratia sp. fermenting pyruvate followed by hydrogenotrophic N2O reduction by a Desulfosporosinus sp. Integrated omics and physiological characterization revealed interspecies nutritional interactions, with the pyruvate fermenting Serratia sp. supplying amino acids as essential growth factors to the N2O-reducing Desulfosporosinus sp. Thus, we demonstrate growth-linked N2O reduction between pH 4.5 and 6, highlighting microbial N2O reduction potential in acidic soils.


Assuntos
Óxido Nitroso , Serratia , Microbiologia do Solo , Óxido Nitroso/metabolismo , Concentração de Íons de Hidrogênio , Serratia/metabolismo , Serratia/genética , Oxirredução , Solo/química , Fermentação , Técnicas de Cocultura , Ácido Pirúvico/metabolismo , Oxirredutases/metabolismo , Oxirredutases/genética , Nitrogênio/metabolismo
8.
Nat Commun ; 15(1): 4158, 2024 May 16.
Artigo em Inglês | MEDLINE | ID: mdl-38755143

RESUMO

Photosynthetic organisms, fungi, and animals comprise distinct pathways for vitamin C biosynthesis. Besides this diversity, the final biosynthetic step consistently involves an oxidation reaction carried out by the aldonolactone oxidoreductases. Here, we study the origin and evolution of the diversified activities and substrate preferences featured by these flavoenzymes using molecular phylogeny, kinetics, mutagenesis, and crystallographic experiments. We find clear evidence that they share a common ancestor. A flavin-interacting amino acid modulates the reactivity with the electron acceptors, including oxygen, and determines whether an enzyme functions as an oxidase or a dehydrogenase. We show that a few side chains in the catalytic cavity impart the reaction stereoselectivity. Ancestral sequence reconstruction outlines how these critical positions were affixed to specific amino acids along the evolution of the major eukaryotic clades. During Eukarya evolution, the aldonolactone oxidoreductases adapted to the varying metabolic demands while retaining their overarching vitamin C-generating function.


Assuntos
Ácido Ascórbico , Evolução Molecular , Filogenia , Ácido Ascórbico/biossíntese , Ácido Ascórbico/metabolismo , Cinética , Oxirredutases/metabolismo , Oxirredutases/genética , Oxirredutases/química , Cristalografia por Raios X , Oxirredução , Animais , Domínio Catalítico , Especificidade por Substrato , Modelos Moleculares
9.
Anal Chim Acta ; 1308: 342664, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38740454

RESUMO

Nanozymes is a kind of nanomaterials with enzyme catalytic properties. Compared with natural enzymes, nanozymes merge the advantages of both nanomaterials and natural enzymes, which is highly important in applications such as biosensing, clinical diagnosis, and food inspection. In this study, we prepared ß-MnOOH hexagonal nanoflakes with a high oxygen vacancy ratio by utilizing SeO2 as a sacrificial agent. The defect-rich MnOOH hexagonal nanoflakes demonstrated excellent oxidase-like activity, catalyzing the oxidation substrate in the presence of O2, thereby rapidly triggering a color reaction. Consequently, a colorimetric sensing platform was constructed to assess the total antioxidant capacity in commercial beverages. The strategy of introducing defects in situ holds great significance for the synthesis of a series of high-performance metal oxide nanozymes, driving the development of faster and more efficient biosensing and analysis methods.


Assuntos
Antioxidantes , Compostos de Manganês , Óxidos , Óxidos/química , Antioxidantes/química , Antioxidantes/metabolismo , Antioxidantes/análise , Compostos de Manganês/química , Colorimetria , Oxirredutases/química , Oxirredutases/metabolismo , Oxirredução , Nanoestruturas/química , Catálise
10.
J Biosci ; 492024.
Artigo em Inglês | MEDLINE | ID: mdl-38726824

RESUMO

Mitochondrial alternative oxidase (AOX) is an important protein that can help in regulating reactive oxygen species and nitric oxide in plants. The role of AOX in regulation of nitro-oxidative stress in chickpea is not known. Using germinating chickpea as a model system, we investigated the role of AOX in nitro-oxidative stress tolerance. NaCl treatment was used as an inducer of nitro-oxidative stress. Treatment of germinating seeds with 150 mM NaCl led to reduced germination and radicle growth. The AOX inhibitor SHAM caused further inhibition of germination, and the AOX inducer pyruvate improved growth of the radicle under NaCl stress. Isolated mitochondria from germinated seeds under salt stress not only increased AOX capacity but also enhanced AOX protein expression. Measurement of superoxide levels revealed that AOX inhibition by SHAM can enhance superoxide levels, whereas the AOX inducer pyruvate reduced superoxide levels. Measurement of NO by gas phase chemiluminescence revealed enhanced NO generation in response to NaCl treatment. Upon NaCl treatment there was enhanced tyrosine nitration, which is an indicator of nitrosative stress response. Taken together, our results revealed that AOX induced under salinity stress in germinating chickpea can help in mitigating nitro-oxidative stress, thereby improving germination.


Assuntos
Cicer , Germinação , Mitocôndrias , Proteínas Mitocondriais , Óxido Nítrico , Estresse Oxidativo , Oxirredutases , Proteínas de Plantas , Superóxidos , Cicer/crescimento & desenvolvimento , Cicer/efeitos dos fármacos , Cicer/metabolismo , Proteínas de Plantas/metabolismo , Germinação/efeitos dos fármacos , Proteínas Mitocondriais/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Óxido Nítrico/metabolismo , Oxirredutases/metabolismo , Superóxidos/metabolismo , Sementes/crescimento & desenvolvimento , Sementes/efeitos dos fármacos , Sementes/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Cloreto de Sódio/farmacologia , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Ácido Pirúvico/metabolismo
11.
Nat Commun ; 15(1): 3802, 2024 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-38714719

RESUMO

The interaction between nuclear receptor coactivator 4 (NCOA4) and the iron storage protein ferritin is a crucial component of cellular iron homeostasis. The binding of NCOA4 to the FTH1 subunits of ferritin initiates ferritinophagy-a ferritin-specific autophagic pathway leading to the release of the iron stored inside ferritin. The dysregulation of NCOA4 is associated with several diseases, including neurodegenerative disorders and cancer, highlighting the NCOA4-ferritin interface as a prime target for drug development. Here, we present the cryo-EM structure of the NCOA4-FTH1 interface, resolving 16 amino acids of NCOA4 that are crucial for the interaction. The characterization of mutants, designed to modulate the NCOA4-FTH1 interaction, is used to validate the significance of the different features of the binding site. Our results explain the role of the large solvent-exposed hydrophobic patch found on the surface of FTH1 and pave the way for the rational development of ferritinophagy modulators.


Assuntos
Microscopia Crioeletrônica , Ferritinas , Coativadores de Receptor Nuclear , Ferritinas/metabolismo , Ferritinas/química , Ferritinas/genética , Humanos , Coativadores de Receptor Nuclear/metabolismo , Coativadores de Receptor Nuclear/química , Coativadores de Receptor Nuclear/genética , Ligação Proteica , Sítios de Ligação , Ferro/metabolismo , Autofagia , Modelos Moleculares , Células HEK293 , Oxirredutases/metabolismo , Oxirredutases/química , Oxirredutases/genética , Proteólise , Mutação
12.
Cell Death Dis ; 15(5): 329, 2024 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-38740757

RESUMO

Iron is crucial for cell DNA synthesis and repair, but an excess of free iron can lead to oxidative stress and subsequent cell death. Although several studies suggest that cancer cells display characteristics of 'Iron addiction', an ongoing debate surrounds the question of whether iron can influence the malignant properties of ovarian cancer. In the current study, we initially found iron levels increase during spheroid formation. Furthermore, iron supplementation can promote cancer cell survival, cancer spheroid growth, and migration; vice versa, iron chelators inhibit this process. Notably, iron reduces the sensitivity of ovarian cancer cells to platinum as well. Mechanistically, iron downregulates DNA homologous recombination (HR) inhibitor polymerase theta (POLQ) and relieves its antagonism against the HR repair enzyme RAD51, thereby promoting DNA damage repair to resist chemotherapy-induced damage. Additionally, iron tightly regulated by ferritin (FTH1/FTL) which is indispensable for iron-triggered DNA repair. Finally, we discovered that iron chelators combined with platinum exhibit a synergistic inhibitory effect on ovarian cancer in vitro and in vivo. Our findings affirm the pro-cancer role of iron in ovarian cancer and reveal that iron advances platinum resistance by promoting DNA damage repair through FTH1/FTL/POLQ/RAD51 pathway. Our findings highlight the significance of iron depletion therapy, revealing a promising avenue for advancing ovarian cancer treatment.


Assuntos
Reparo do DNA , Resistencia a Medicamentos Antineoplásicos , Ferro , Neoplasias Ovarianas , Rad51 Recombinase , Feminino , Humanos , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Neoplasias Ovarianas/genética , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Reparo do DNA/efeitos dos fármacos , Ferro/metabolismo , Linhagem Celular Tumoral , Rad51 Recombinase/metabolismo , Animais , Ferritinas/metabolismo , Camundongos , Platina/farmacologia , Platina/uso terapêutico , Camundongos Nus , Oxirredutases/metabolismo
13.
Nat Commun ; 15(1): 4226, 2024 May 18.
Artigo em Inglês | MEDLINE | ID: mdl-38762502

RESUMO

Aerobic methanotrophic bacteria are considered strict aerobes but are often highly abundant in hypoxic and even anoxic environments. Despite possessing denitrification genes, it remains to be verified whether denitrification contributes to their growth. Here, we show that acidophilic methanotrophs can respire nitrous oxide (N2O) and grow anaerobically on diverse non-methane substrates, including methanol, C-C substrates, and hydrogen. We study two strains that possess N2O reductase genes: Methylocella tundrae T4 and Methylacidiphilum caldifontis IT6. We show that N2O respiration supports growth of Methylacidiphilum caldifontis at an extremely acidic pH of 2.0, exceeding the known physiological pH limits for microbial N2O consumption. Methylocella tundrae simultaneously consumes N2O and CH4 in suboxic conditions, indicating robustness of its N2O reductase activity in the presence of O2. Furthermore, in O2-limiting conditions, the amount of CH4 oxidized per O2 reduced increases when N2O is added, indicating that Methylocella tundrae can direct more O2 towards methane monooxygenase. Thus, our results demonstrate that some methanotrophs can respire N2O independently or simultaneously with O2, which may facilitate their growth and survival in dynamic environments. Such metabolic capability enables these bacteria to simultaneously reduce the release of the key greenhouse gases CO2, CH4, and N2O.


Assuntos
Metano , Óxido Nitroso , Óxido Nitroso/metabolismo , Metano/metabolismo , Concentração de Íons de Hidrogênio , Oxirredutases/metabolismo , Oxirredutases/genética , Oxigênio/metabolismo , Oxirredução , Anaerobiose , Metanol/metabolismo , Hidrogênio/metabolismo , Oxigenases/metabolismo , Oxigenases/genética
14.
BMC Plant Biol ; 24(1): 232, 2024 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-38561659

RESUMO

BACKGROUND: Chrysanthemum, one of the four major cut flowers all over the world, is very sensitive to salinity during cultivation. DNA binding with one finger (DOF) transcription factors play important roles in biological processes in plants. The response mechanism of CmDOF18 from chrysanthemum to salt stress remains unclear. RESULTS: In this study, CmDOF18 was cloned from Chrysanthemum morifolium, and its expression was induced by salinity stress. The gene encodes a 291-amino acid protein with a typical DOF domain. CmDOF18 was localized to the nucleus in onion epidermal cells and showed transcriptional activation in yeast. CmDOF18 transgenic plants were generated to identify the role of this gene in resistance to salinity treatment. Chrysanthemum plants overexpressing CmDOF18 were more resistant to salinity stress than wild-type plants. Under salinity stress, the malondialdehyde content and leaf electrolyte conductivity in CmDOF18-overexpressing transgenic plants were lower than those in wild-type plants, while the proline content, chlorophyll content, superoxide dismutase activity and peroxidase activity were higher than those in wild-type plants. The opposite findings were observed in gene-silenced plants compared with wild-type plants. The gene expression levels of oxidoreductase increased in CmDOF18-overexpressing transgenic plants but decreased in CmDOF18-SRDX gene-silenced transgenic plants. CONCLUSION: In summary, we analyzed the function of CmDOF18 from chrysanthemum, which may regulate salinity stress in plants, possibly due to its role in the regulation of oxidoreductase.


Assuntos
Chrysanthemum , Oxirredutases , Oxirredutases/metabolismo , Tolerância ao Sal/genética , Chrysanthemum/genética , Chrysanthemum/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Saccharomyces cerevisiae/metabolismo , Salinidade , Regulação da Expressão Gênica de Plantas , Estresse Fisiológico/genética
15.
Environ Sci Technol ; 58(16): 7056-7065, 2024 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-38608141

RESUMO

The sources and sinks of nitrous oxide, as control emissions to the atmosphere, are generally poorly constrained for most environmental systems. Initial depth-resolved analysis of nitrous oxide flux from observation wells and the proximal surface within a nitrate contaminated aquifer system revealed high subsurface production but little escape from the surface. To better understand the environmental controls of production and emission at this site, we used a combination of isotopic, geochemical, and molecular analyses to show that chemodenitrification and bacterial denitrification are major sources of nitrous oxide in this subsurface, where low DO, low pH, and high nitrate are correlated with significant nitrous oxide production. Depth-resolved metagenomes showed that consumption of nitrous oxide near the surface was correlated with an enrichment of Clade II nitrous oxide reducers, consistent with a growing appreciation of their importance in controlling release of nitrous oxide to the atmosphere. Our work also provides evidence for the reduction of nitrous oxide at a pH of 4, well below the generally accepted limit of pH 5.


Assuntos
Óxido Nitroso , Óxido Nitroso/metabolismo , Bactérias/metabolismo , Oxirredutases/metabolismo , Desnitrificação
16.
Food Chem ; 448: 139170, 2024 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-38579558

RESUMO

Current nanozyme applications rely heavily on peroxidase-like nanozymes and are limited to a specific temperature range, despite notable advancements in nanozyme development. In this work, we designed novel Mn-based metal organic frameworks (UoZ-4), with excellent oxidase mimic activity towards common substrates. UoZ-4 showed excellent oxidase-like activity (with Km 0.072 mM) in a wide range of temperature, from 10 °C to 100 °C with almost no activity loss, making it a very strong candidate for psychrophilic and thermophilic applications. Ascorbic acid, cysteine, and glutathione could quench the appearance of the blue color of oxTMB, led us to design a visual-based sensing platform for detection of total antioxidant capacity (TAC) in cold, mild and hot conditions. The visual mode successfully assessed TAC in citrus fruits with satisfactory recovery and precisions. Cold/hot adapted and magnetic property will broaden the horizon of nanozyme applications and breaks the notion of the temperature limitation of enzymes.


Assuntos
Antioxidantes , Citrus , Frutas , Manganês , Estruturas Metalorgânicas , Oxirredutases , Temperatura , Citrus/química , Citrus/metabolismo , Antioxidantes/metabolismo , Antioxidantes/química , Antioxidantes/análise , Frutas/química , Frutas/metabolismo , Manganês/metabolismo , Manganês/química , Manganês/análise , Estruturas Metalorgânicas/química , Oxirredutases/metabolismo , Oxirredutases/química
17.
Appl Environ Microbiol ; 90(4): e0014624, 2024 Apr 17.
Artigo em Inglês | MEDLINE | ID: mdl-38557120

RESUMO

The metal-resistant bacterium Cupriavidus metallidurans occurs in metal-rich environments. In auriferous soils, the bacterium is challenged by a mixture of copper ions and gold complexes, which exert synergistic toxicity. The previously used, self-made Au(III) solution caused a synergistic toxicity of copper and gold that was based on the inhibition of the CupA-mediated efflux of cytoplasmic Cu(I) by Au(I) in this cellular compartment. In this publication, the response of the bacterium to gold and copper was investigated by using a commercially available Au(III) solution instead of the self-made solution. The new solution was five times more toxic than the previously used one. Increased toxicity was accompanied by greater accumulation of gold atoms by the cells. The contribution of copper resistance determinants to the commercially available Au(III) solution and synergistic gold-copper toxicity was studied using single- and multiple-deletion mutants. The commercially available Au(III) solution inhibited periplasmic Cu(I) homeostasis, which is required for the allocation of copper ions to copper-dependent proteins in this compartment. The presence of the gene for the periplasmic Cu(I) and Au(I) oxidase, CopA, decreased the cellular copper and gold content. Transcriptional reporter gene fusions showed that up-regulation of gig, encoding a minor contributor to copper resistance, was strictly glutathione dependent. Glutathione was also required to resist synergistic gold-copper toxicity. The new data indicated a second layer of synergistic copper-gold toxicity caused by the commercial Au(III) solution, inhibition of the periplasmic copper homeostasis in addition to the cytoplasmic one.IMPORTANCEWhen living in auriferous soils, Cupriavidus metallidurans is not only confronted with synergistic toxicity of copper ions and gold complexes but also by different gold species. A previously used gold solution made by using aqua regia resulted in the formation of periplasmic gold nanoparticles, and the cells were protected against gold toxicity by the periplasmic Cu(I) and Au(I) oxidase CopA. To understand the role of different gold species in the environment, another Au(III) solution was commercially acquired. This compound was more toxic due to a higher accumulation of gold atoms by the cells and inhibition of periplasmic Cu(I) homeostasis. Thus, the geo-biochemical conditions might influence Au(III) speciation. The resulting Au(III) species may subsequently interact in different ways with C. metallidurans and its copper homeostasis system in the cytoplasm and periplasm. This study reveals that the geochemical conditions may decide whether bacteria are able to form gold nanoparticles or not.


Assuntos
Cupriavidus , Nanopartículas Metálicas , Cobre/metabolismo , Ouro/toxicidade , Ouro/metabolismo , Nanopartículas Metálicas/toxicidade , Nanopartículas Metálicas/química , Cupriavidus/genética , Cupriavidus/metabolismo , Proteínas de Bactérias/metabolismo , Íons/metabolismo , Solo , Glutationa/metabolismo , Oxirredutases/metabolismo
18.
Sci Total Environ ; 930: 172695, 2024 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-38663613

RESUMO

General control non-derepressible-2 (GCN2) is widely expressed in eukaryotes and responds to biotic and abiotic stressors. However, the precise function and mechanism of action of GCN2 in response to cadmium (Cd) stress in Nicotiana tabacum L. (tobacco) remains unclear. We investigated the role of NtGCN2 in Cd tolerance and explored the mechanism by which NtGCN2 responds to Cd stress in tobacco by exposing NtGCN2 transgenic tobacco lines to different concentrations of CdCl2. NtGCN2 was activated under 50 µmol·L-1 CdCl2 stress and enhanced the Cd tolerance and photosynthetic capacities of tobacco by increasing chlorophyll content and antioxidant capacity by upregulating NtSOD, NtPOD, and NtCAT expression and corresponding enzyme activities and decreasing malondialdehyde and O2·- contents. NtGCN2 enhanced the osmoregulatory capacity of tobacco by elevating proline (Pro) and soluble sugar contents and maintaining low levels of relative conductivity. Finally, NtGCN2 enhanced Cd tolerance in tobacco by reducing Cd uptake and translocation, promoting Cd efflux, and regulating Cd subcellular distribution. In conclusion, NtGCN2 improves the tolerance of tobacco to Cd through a series of mechanisms, namely, increasing antioxidant, photosynthetic, and osmoregulation capacities and regulating Cd uptake, translocation, efflux, and subcellular distribution. This study provides a scientific basis for further exploration of the role of NtGCN2 in plant responses to Cd stress and enhancement of the Cd stress signaling network in tobacco.


Assuntos
Cádmio , Resistência a Medicamentos , Nicotiana , Proteínas de Plantas , Cádmio/toxicidade , Cádmio/metabolismo , Nicotiana/fisiologia , Nicotiana/metabolismo , Fotossíntese/efeitos dos fármacos , Fotossíntese/genética , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genética , Poluentes do Solo/metabolismo , Poluentes do Solo/toxicidade , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , Clorofila/metabolismo , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/genética , Folhas de Planta/metabolismo , Resistência a Medicamentos/genética , Oxirredutases/genética , Oxirredutases/metabolismo , Ativação Enzimática/genética , Osmorregulação/genética , Espaço Intracelular/metabolismo
19.
Chemosphere ; 357: 142029, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38626812

RESUMO

The application of herbicides in soil has been noted for its detrimental effect on the soil microbial community, crucial for various biochemical processes. This study provides a comprehensive assessment of the impact of butisanstar and clopyralid herbicides, both individually and in combination at different dosage (recommended field dose (RFD), ½, 2 and 5-times RFD). The assessment focuses on soil basal respiration (SBR), cumulative microbial respiration (CMR), and the activities dehydrogenase (DH), catalase (CAT), urease, acid and alkaline phosphatases (Ac-P and Alk-P) enzymes, along with their variations on days 10, 30, 60, and 90 post-herbicide application. Results indicate that, although herbicides, even at lower doses of RFD, demonstrate inhibitory effects on DH, CAT, and microbial respiration, they paradoxically lead to a significant enhancement in urease and phosphatase activities, even at higher doses. The inhibitory/enhancing intensity varies based on herbicide type, incubation period, and dosage. Co-application of herbicides manifests synergistic effects compared to individual applications. The most notable inhibitory effects on DH, CAT, and SBR are observed on the 30th day, coinciding with the highest activities of urease and phosphatases on the same day. The persistent inability to restore respiration and enzyme activities to initial soil (control) levels emphasizes the lasting adverse and inhibitory effects of herbicides, especially clopyralid, over the long term. It becomes apparent that soil microorganisms require an extended duration to decompose and acclimate to the presence of herbicides. Consequently, these agrochemical compounds pose a potential risk to crucial biochemical processes, such as nutrient cycling, ultimately impacting crop production.


Assuntos
Herbicidas , Microbiologia do Solo , Poluentes do Solo , Solo , Herbicidas/toxicidade , Poluentes do Solo/toxicidade , Solo/química , Catalase/metabolismo , Ecotoxicologia , Urease/metabolismo , Oxirredutases/metabolismo
20.
ACS Infect Dis ; 10(5): 1739-1752, 2024 May 10.
Artigo em Inglês | MEDLINE | ID: mdl-38647213

RESUMO

Reverse analogs of the phosphonohydroxamic acid antibiotic fosmidomycin are potent inhibitors of the nonmevalonate isoprenoid biosynthesis enzyme 1-deoxy-d-xylulose 5-phosphate reductoisomerase (DXR, IspC) of Plasmodium falciparum. Some novel analogs with large phenylalkyl substituents at the hydroxamic acid nitrogen exhibit nanomolar PfDXR inhibition and potent in vitro growth inhibition of P. falciparum parasites coupled with good parasite selectivity. X-ray crystallographic studies demonstrated that the N-phenylpropyl substituent of the newly developed lead compound 13e is accommodated in a subpocket within the DXR catalytic domain but does not reach the NADPH binding pocket of the N-terminal domain. As shown for reverse carba and thia analogs, PfDXR selectively binds the S-enantiomer of the new lead compound. In addition, some representatives of the novel inhibitor subclass are nanomolar Escherichia coli DXR inhibitors, whereas the inhibition of Mycobacterium tuberculosis DXR is considerably weaker.


Assuntos
Aldose-Cetose Isomerases , Antimaláricos , Fosfomicina , Ácidos Hidroxâmicos , Complexos Multienzimáticos , Plasmodium falciparum , Fosfomicina/farmacologia , Fosfomicina/análogos & derivados , Fosfomicina/química , Aldose-Cetose Isomerases/antagonistas & inibidores , Aldose-Cetose Isomerases/metabolismo , Aldose-Cetose Isomerases/química , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/enzimologia , Ácidos Hidroxâmicos/farmacologia , Ácidos Hidroxâmicos/química , Antimaláricos/farmacologia , Antimaláricos/química , Complexos Multienzimáticos/antagonistas & inibidores , Complexos Multienzimáticos/metabolismo , Complexos Multienzimáticos/química , Cristalografia por Raios X , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/química , Inibidores Enzimáticos/síntese química , Relação Estrutura-Atividade , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/enzimologia , Modelos Moleculares , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/enzimologia , Domínio Catalítico , Oxirredutases/antagonistas & inibidores , Oxirredutases/metabolismo
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