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1.
Commun Biol ; 4(1): 841, 2021 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-34230602

RESUMO

Characterizing protein-protein interactions (PPIs) is an effective method to help explore protein function. Here, through integrating a newly identified split human Rhinovirus 3 C (HRV 3 C) protease, super-folder GFP (sfGFP), and ClpXP-SsrA protein degradation machinery, we developed a fluorescence-assisted single-cell methodology (split protease-E. coli ClpXP (SPEC)) to explore protein-protein interactions for both eukaryotic and prokaryotic species in E. coli cells. We firstly identified a highly efficient split HRV 3 C protease with high re-assembly ability and then incorporated it into the SPEC method. The SPEC method could convert the cellular protein-protein interaction to quantitative fluorescence signals through a split HRV 3 C protease-mediated proteolytic reaction with high efficiency and broad temperature adaptability. Using SPEC method, we explored the interactions among effectors of representative type I-E and I-F CRISPR/Cas complexes, which combining with subsequent studies of Cas3 mutations conferred further understanding of the functions and structures of CRISPR/Cas complexes.


Assuntos
Endopeptidase Clp/metabolismo , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Mapas de Interação de Proteínas , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sistemas CRISPR-Cas , Endopeptidase Clp/genética , Enterovirus/enzimologia , Enterovirus/genética , Escherichia coli/genética , Proteínas de Escherichia coli/genética , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Histidina Quinase/genética , Histidina Quinase/metabolismo , Humanos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Proteólise , Serina Endopeptidases/química , Serina Endopeptidases/genética , Serina Endopeptidases/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismo
2.
Appl Microbiol Biotechnol ; 105(12): 5077-5086, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-34106311

RESUMO

Due to its robustness to environmental stresses and fast growth, Synechococcus elongatus UTEX2973 is developed as a new model for researches on cyanobacterial molecular biology and biotechnology. However, systematic genetic modifications of S. elongatus UTEX2973 were hindered by the lack of effective genetic manipulation tools, especially available counter-selection markers. Here, six synthetic counter-selection markers (SCOMs) were assembled by fusing six toxin genes from either Escherichia coli or cyanobacteria with a theophylline-inducible promoter. The SCOMs containing SYNPCC7002_G0085 from Synechococcus sp. PCC7002 or mazF from E. coli were proved to be inducible by theophylline in S. elongatus UTEX2973. By using the mazF-based SCOM, the neutral locus 1 and 23 small regulatory RNAs were completely deleted from the genome of S. elongatus UTEX2973 after one round of selection with both kanamycin and theophylline. The genetic tools developed in this work will facilitate future researches on molecular genetics and synthetic biology in S. elongatus UTEX2973. KEY POINTS: • Two inducible counter-selection markers are lethal to S. elongatus UTEX2973. • The counter-selection marker benefits the gene targeting in S. elongatus UTEX2973. • Twentry-three small regulatory RNAs were fully deleted via the novel gene targeting method.


Assuntos
Proteínas de Escherichia coli , Synechococcus , Proteínas de Ligação a DNA , Endorribonucleases , Escherichia coli , Regiões Promotoras Genéticas , Synechococcus/genética
3.
J Periodontal Res ; 56(5): 837-847, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34173676

RESUMO

Curcumin is the main active ingredient of turmeric, which has a wide range of pharmacological effects, including antitumor, antibacterial, anti-inflammatory, anti-oxidation, immune regulation, and so on. Periodontitis is a prevalent oral inflammatory disease caused by a variety of factors. In recent years, many studies have shown that curcumin has a potential role on the treatment of periodontitis. Curcumin has been used in research related to the treatment of periodontitis in the form of solution, chip, gel, and capsule. Combined with other periodontitis treatment methods, such as scaling and root planing (SRP) and photodynamic therapy (PDT), can enhance curcumin's efficacy in treating periodontitis. In addition to natural curcumin, chemically modified curcumin, such as 4-phenylaminocarbonyl bis-demethoxy curcumin (CMC 2.24) and 4-methoxycarbonyl curcumin (CMC 2.5), have also been used in animal models of periodontitis. Here, this paper reviews the research progress of curcumin on the treatment of periodontitis and its related mechanisms.


Assuntos
Curcumina , Periodontite , Animais , Anti-Inflamatórios/uso terapêutico , Curcumina/uso terapêutico , Raspagem Dentária , Periodontite/tratamento farmacológico , Aplainamento Radicular
4.
Sheng Wu Gong Cheng Xue Bao ; 37(3): 874-910, 2021 Mar 25.
Artigo em Chinês | MEDLINE | ID: mdl-33783156

RESUMO

The development and implement of microbial chassis cells can provide excellent cell factories for diverse industrial applications, which help achieve the goal of environmental protection and sustainable bioeconomy. The synthetic biology strategy of Design-Build-Test-Learn (DBTL) plays a crucial role on rational and/or semi-rational construction or modification of chassis cells to achieve the goals of "Building to Understand" and "Building for Applications". In this review, we briefly comment on the technical development of the DBTL cycle and the research progress of a few model microorganisms. We mainly focuse on non-model bacterial cell factories with potential industrial applications, which possess unique physiological and biochemical characteristics, capabilities of utilizing one-carbon compounds or of producing platform compounds efficiently. We also propose strategies for the efficient and effective construction and application of synthetic microbial cell factories securely in the synthetic biology era, which are to discover and integrate the advantages of model and non-model industrial microorganisms, to develop and deploy intelligent automated equipment for cost-effective high-throughput screening and characterization of chassis cells as well as big-data platforms for storing, retrieving, analyzing, simulating, integrating, and visualizing omics datasets at both molecular and phenotypic levels, so that we can build both high-quality digital cell models and optimized chassis cells to guide the rational design and construction of microbial cell factories for diverse industrial applications.


Assuntos
Engenharia Metabólica , Biologia Sintética , Bactérias/genética
5.
Cancer Gene Ther ; 2021 Feb 02.
Artigo em Inglês | MEDLINE | ID: mdl-33531646

RESUMO

Increasing evidence proved the abnormal expression of long non-coding RNAs (lncRNAs) in various human malignancies, including oral squamous cell carcinoma (OSCC). Nevertheless, limited explorations concern the role of lncRNA small nucleolar RNA host gene 17 (SNHG17) in OSCC. Herein, SNHG17 was disclosed to be remarkably upregulated in OSCC cell lines and promoted OSCC cell growth. Further mechanistic studies, including DNA/RNA pull down, RIP, ChIP, and luciferase reporter gene assays, were conducted. It was confirmed that Wnt/ß-catenin signaling pathway was involved in the SNHG17-mediated OSCC cell growth. Moreover, E74 like ETS transcription factor 1 (ELF1) was identified as the transcription activator of CTNNB1 (ß-catenin mRNA) in OSCC. Inspired by competing for endogenous RNAs (ceRNAs) network, we were pleasantly surprised to find that SNHG17 and ELF1 functioned as ceRNAs in OSCC via competitively binding to microRNA-384 (miR-384). By using rescue assays, we revealed that SNHG17 facilitated OSCC cell growth through modulating miR-384/ELF1 axis. Importantly, we certified that ELF1 was indispensable for SNHG17-affected OSCC progression. Collectively, it can be concluded that SNHG17/miR-384/ELF1 axis contributed to OSCC cell growth via promoting CTNNB1 expression, thus activating Wnt/ß-catenin signaling pathway.

6.
Front Microbiol ; 11: 597454, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33250885

RESUMO

We isolated an aromatic strain of yeast (M2013310) from chili sauce. Assembly, annotation, and phylogenetic analysis based on genome sequencing, identified M2013310 as an allodiploid yeast that was closely related to Zygosaccharomyces rouxii. During fermentation, M2013310, produced an aromatic alcohol with a rose-honey scent; gas chromatography tandem mass spectrometry identified this alcohol as 2-phenylethanol. The concentration of 2-phenylethanol reached 3.8 mg/L, 1.79 g/L, and 3.58 g/L, in M3 (NH4 +), M3 (NH4 + + Phe), and M3 (Phe) culture media, after 72 h of fermentation, respectively. The mRNA expression levels of ARO8 encoding aromatic aminotransferases I and ARO10 encoding phenylpyruvate decarboxylase by M2013310 in M3 (Phe) were the lowest of the three different forms of media tested. These results indicated that M2013310 can synthesize 2-phenylethanol via the Shikimate or Ehrlich pathways and the production of 2-phenylethanol may be significantly improved by the over-expression of these two genes. Our research identified a promising strain of yeast (M2013310) that could be used to improve the production of 2-phenylethanol.

7.
Artigo em Inglês | MEDLINE | ID: mdl-32984271

RESUMO

Zymomonas mobilis is a model ethanologenic bacterium for diverse biochemical production. Rich medium (RM) is a complex medium that is routinely used to cultivate Z. mobilis, which contains carbon sources such as glucose, nitrogen sources such as yeast extract (YE), and KH2PO4. Glucose consumption and cell growth of Z. mobilis is usually coupled during ethanol fermentation. However, sometimes glucose was not consumed during the exponential growth phase, and it took extended time for cells to consume glucose and produce ethanol, which eventually reduced the ethanol productivity. In this study, the effects of different nitrogen sources, as well as the supplementation of an additional nitrogen source into RM and minimal medium (MM), on cell growth and glucose consumption of Z. mobilis were investigated to understand the uncoupled cell growth and glucose consumption. Our results indicated that nitrogen sources such as YE from different companies affected cell growth, glucose utilization, and ethanol production. We also quantified the concentrations of major ion elements in different nitrogen sources using the quantitative analytic approach of Inductively Coupled Plasma Optical Emission Spectroscopy (ICP-OES), and demonstrated that magnesium ion in the media affected cell growth, glucose consumption, and ethanol production. The effect of magnesium on gene expression was further investigated using RNA-Seq transcriptomics. Our results indicated that the lack of Mg2+ triggered stress responses, and the expression of genes involved in energy metabolism was reduced. Our work thus demonstrated that Mg2+concentration in nitrogen sources is essential for vigorous cell growth and ethanol fermentation, and the difference of Mg2+concentration in different YE is one of the major factors affecting the coupled cell growth, glucose consumption and ethanol fermentation in Z. mobilis. We also revealed that genes responsive for Mg2+ deficiency in the medium were majorly related to stress responses and energy conservation. The importance of magnesium on cell growth and ethanol fermentation suggests that metal ions should become one of the parameters for monitoring the quality of commercial nitrogen sources and optimizing microbial culture medium.

8.
Biomed Mater ; 15(5): 055012, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32857733

RESUMO

Titanium (Ti) and its alloys are widely used in dental implants due to their favorable mechanical properties and biocompatibility. Surface characteristics, including physical and chemical properties, are crucial factors to enhance the osteogenesis performance of Ti. The aim of this study is to evaluate amino group surface modification to facilitate the osteogenic potential and bone repair of dental implants both in vitro and in vivo. Herein, amino group-modified Ti surfaces were constructed via the plasma-enhanced chemical vapor deposition (PECVD) technique with an allylamine monomer. The adhesion, proliferation, alkaline phosphate activity and osteogenesis-related genetic expression of MG-63 cells on the surfaces were performed in vitro and presented a significant increase in amino group-modified Ti compared with that in Ti. The in vivo study in miniature pigs was evaluated through micro-computed tomography analysis and histological evaluation, which exhibited enhanced new bone formation in amino group-modified Ti compared with that in Ti after implantation for 4, 8 and 12 weeks. Consequently, amino group surface modification with the PECVD technique may provide a promising modification method to enhance the osteogenesis performance of Ti implants.

9.
Biotechnol Biofuels ; 13: 144, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32817760

RESUMO

Background: Acid pretreatment is a common strategy used to break down the hemicellulose component of the lignocellulosic biomass to release pentoses, and a subsequent enzymatic hydrolysis step is usually applied to release hexoses from the cellulose. The hydrolysate after pretreatment and enzymatic hydrolysis containing both hexoses and pentoses can then be used as substrates for biochemical production. However, the acid-pretreated liquor can also be directly used as the substrate for microbial fermentation, which has an acidic pH and contains inhibitory compounds generated during pretreatment. Although the natural ethanologenic bacterium Zymomonas mobilis can grow in a broad range of pH 3.5 ~ 7.5, cell growth and ethanol fermentation are still affected under acidic-pH conditions below pH 4.0. Results: In this study, adaptive laboratory evolution (ALE) strategy was applied to adapt Z. mobilis under acidic-pH conditions. Two mutant strains named 3.6M and 3.5M with enhanced acidic pH tolerance were selected and confirmed, of which 3.5M grew better than ZM4 but worse than 3.6M in acidic-pH conditions that is served as a reference strain between 3.6M and ZM4 to help unravel the acidic-pH tolerance mechanism. Mutant strains 3.5M and 3.6M exhibited 50 ~ 130% enhancement on growth rate, 4 ~ 9 h reduction on fermentation time to consume glucose, and 20 ~ 63% improvement on ethanol productivity than wild-type ZM4 at pH 3.8. Next-generation sequencing (NGS)-based whole-genome resequencing (WGR) and RNA-Seq technologies were applied to unravel the acidic-pH tolerance mechanism of mutant strains. WGR result indicated that compared to wild-type ZM4, 3.5M and 3.6M have seven and five single nucleotide polymorphisms (SNPs), respectively, among which four are shared in common. Additionally, RNA-Seq result showed that the upregulation of genes involved in glycolysis and the downregulation of flagellar and mobility related genes would help generate and redistribute cellular energy to resist acidic pH while keeping normal biological processes in Z. mobilis. Moreover, genes involved in RND efflux pump, ATP-binding cassette (ABC) transporter, proton consumption, and alkaline metabolite production were significantly upregulated in mutants under the acidic-pH condition compared with ZM4, which could help maintain the pH homeostasis in mutant strains for acidic-pH resistance. Furthermore, our results demonstrated that in mutant 3.6M, genes encoding F1F0 ATPase to pump excess protons out of cells were upregulated under pH 3.8 compared to pH 6.2. This difference might help mutant 3.6M manage acidic conditions better than ZM4 and 3.5M. A few gene targets were then selected for genetics study to explore their role in acidic pH tolerance, and our results demonstrated that the expression of two operons in the shuttle plasmids, ZMO0956-ZMO0958 encoding cytochrome bc1 complex and ZMO1428-ZMO1432 encoding RND efflux pump, could help Z. mobilis tolerate acidic-pH conditions. Conclusion: An acidic-pH-tolerant mutant 3.6M obtained through this study can be used for commercial bioethanol production under acidic fermentation conditions. In addition, the molecular mechanism of acidic pH tolerance of Z. mobilis was further proposed, which can facilitate future research on rational design of synthetic microorganisms with enhanced tolerance against acidic-pH conditions. Moreover, the strategy developed in this study combining approaches of ALE, genome resequencing, RNA-Seq, and classical genetics study for mutant evolution and characterization can be applied in other industrial microorganisms.

10.
Methods Mol Biol ; 2096: 217-233, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32720157

RESUMO

Although rational design-based metabolic engineering has been applied widely to obtain promising microbial biocatalysts, conventional strategies such as adaptive laboratory evolution (ALE) and mutagenesis are still efficient approaches to improve microorganisms for exceptional features such as a broad spectrum of substrate utilization, robustness of cell growth, as well as high titer, yield, and productivity of the target products. In this chapter, we describe the procedure to generate mutant strains with desired phenotypes using ALE and a new mutagenesis approach of Atmosphere and Room Temperature Plasma (ARTP). In addition, we discuss the methodology to combine next-generation sequencing (NGS)-based genome-resequencing and RNA-Seq transcriptomics approaches to characterize the mutant strains and connect the phenotypes with their corresponding genotypic changes.


Assuntos
Bactérias/genética , Genômica/métodos , Biologia Computacional , DNA Bacteriano/isolamento & purificação , DNA Complementar/genética , Evolução Molecular Direcionada , Genoma Bacteriano , Genótipo , Mutagênese/genética , Mutação/genética , Fenótipo , Gases em Plasma/química , RNA Bacteriano/isolamento & purificação , Padrões de Referência , Temperatura
11.
Appl Microbiol Biotechnol ; 104(17): 7247-7260, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32666183

RESUMO

Pectate lyase treatment can be an alternative strategy of the chemical processing, which causes severe environmental pollution, and has been broadly studied and applied for diverse industrial applications including textile industry, beverage industry, pulp processing, pectic wastewater pretreatment, and oil extraction. This review gave a brief description of the origins, enzymatic characterizations, structure, and applications of pectate lyases (Pels). Most of the reported pectate lyases are originated from microorganisms with a small number of them from plants and animals. Due to the diverse environments that these microorganisms exist, Pels present diversified features, especially for the range of optimal pH and temperature. The diversified biochemical properties of Pels define their applications in different industries, and the applications of alkaline Pels on cotton bioscouring and ramie degumming in textile industry were focused in this review. This review also discussed the perspectives of the development and applications of Pels. KEY POINTS: • The first review on pectate lyase focusing on biotechnological applications. • Origins, features, structures, applications of pectate lyases reviewed. • Applications of alkaline Pels in textile industry demonstrated. • Perspectives on future development and applications of Pels discussed.


Assuntos
Pectinas , Polissacarídeo-Liases , Clonagem Molecular
12.
Mar Drugs ; 18(7)2020 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-32610482

RESUMO

: Astaxanthin from H. pluvialis is an antioxidant and presents a promising application in medicine for human health. The two-stage strategy has been widely adopted to produce astaxanthin by the Haematococcus industry and research community. However, cell death and low astaxanthin productivity have seriously affected the stability of astaxanthin production. This study aims to test the effect of cell transformation strategies on the production of astaxanthin from H. pluvialis and determine the optimal initial biomass density (IBD) in the red stage. The experimental design is divided into two parts, one is the vegetative growth experiment and the other is the stress experiment. The results indicated that: (1) the cell transformation strategy of H. pluvialis can effectively reduce cell death occurred in the red stage and significantly increase the biomass and astaxanthin production. (2) Compared with the control group, the cell mortality rate of the red stage in the treatment group was reduced by up to 81.6%, and the biomass and astaxanthin production was increased by 1.63 times and 2.1 times, respectively. (3) The optimal IBD was determined to be 0.5, and the highest astaxanthin content can reach 38.02 ± 2.40 mg·g-1. Thus, this work sought to give useful information that will lead to an improved understanding of the cost-effective method of cultivation of H. pluvialis for natural astaxanthin. This will be profitable for algal and medicine industry players.


Assuntos
Clorófitas/metabolismo , Microalgas/metabolismo , Biomassa , Técnicas de Cultura de Células/métodos , Clorófitas/crescimento & desenvolvimento , Meios de Cultura , Microbiologia Industrial/métodos , Microalgas/crescimento & desenvolvimento , Xantofilas/metabolismo
13.
Colloids Surf B Biointerfaces ; 193: 111098, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32498001

RESUMO

Polyetheretherketone (PEEK) is considered as a potential dental and orthopedic implant material owing to its favorable thermal and chemical stability, biocompatibility and mechanical properties. However, the inherent bio-inert and inferior osseointegration of PEEK have hampered its clinical application. In addition, carbon fiber is widely used as a filler to reinforce polymers for sturdy composites owing to its high strength, modulus, etc. In the study, carbon fiber reinforced PEEK (CPEEK) composites were fabricated and modified with amino groups by plasma-enhanced chemical vapor deposition surface modification technique. The surface characterization of composites was evaluated by FE-SEM, EDS, AFM, Water contact angle, XPS and FTIR, which revealed that amino groups were successfully incorporated on the modified CPEEK surface and significantly increased the hydrophilicity. In vitro study, cell adhesion, proliferation, ALP activity, ECM mineralization, real-time PCR analysis, and ELISA analysis showed the adhesion, proliferation and osteogenic differentiation of MG-63 cells on the amino group-modified CPEEK surface were higher than the CPEEK, equal to or better than pure titanium. Hence, the results indicated that the amino group-modified CPEEK possessed enhanced bioactivity and osteogenic property, which may be a potential candidate material for dental implants.


Assuntos
Materiais Biocompatíveis/farmacologia , Fibra de Carbono/química , Cetonas/farmacologia , Polietilenoglicóis/farmacologia , Materiais Biocompatíveis/química , Adesão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Humanos , Cetonas/química , Osteoblastos/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Tamanho da Partícula , Polietilenoglicóis/química , Propriedades de Superfície
14.
Front Microbiol ; 11: 634, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32362881

RESUMO

Sustainable production of biofuels and biochemicals has been broadly accepted as a solution to lower carbon dioxide emissions. Besides being used as lubricants or detergents, oleochemicals are also attractive biofuels as they are compatible with existing transport infrastructures. Cyanobacteria are autotrophic prokaryotes possessing photosynthetic abilities with mature genetic manipulation systems. Through the introduction of exogenous or the modification of intrinsic metabolic pathways, cyanobacteria have been engineered to produce various bio-chemicals and biofuels over the past decade. In this review, we specifically summarize recent progress on photosynthetic production of fatty acids, fatty alcohols, fatty alk(a/e)nes, and fatty acid esters by genetically engineered cyanobacteria. We also summarize recent reports on fatty acid and lipid metabolisms of cyanobacteria and provide perspectives for economic cyanobacterial oleochemical production in the future.

15.
Artigo em Inglês | MEDLINE | ID: mdl-32318556

RESUMO

The methane (CH4)/oxygen (O2) gas supply ratios significantly affect the cell growth and metabolic pathways of aerobic obligate methanotrophs. However, few studies have explored the CH4/O2 ratios of the inlet gas, especially for the CH4 concentrations within the explosion range (5∼15% of CH4 in air). This study thoroughly investigated the molecular mechanisms associated with the impact of different CH4/O2 ratios on cell growth of a model type I methanotroph Methylomicrobium buryatense 5GB1 cultured at five different CH4/O2 supply molar ratios from 0.28 to 5.24, corresponding to CH4 content in gas mixture from 5% to 50%, using RNA-Seq transcriptomics approach. In the batch cultivation, the highest growth rate of 0.287 h-1 was achieved when the CH4/O2 supply molar ratio was 0.93 (15% CH4 in air), and it is crucial to keep the availability of carbon and oxygen levels balanced for optimal growth. At this ratio, genes related to methane metabolism, phosphate uptake system, and nitrogen fixation were significantly upregulated. The results indicated that the optimal CH4/O2 ratio prompted cell growth by increasing genes involved in metabolic pathways of carbon, nitrogen and phosphate utilization in M. buryatense 5GB1. Our findings provided an effective gas supply strategy for methanotrophs, which could enhance the production of key intermediates and enzymes to improve the performance of bioconversion processes using CH4 as the only carbon and energy source. This research also helps identify genes associated with the optimal CH4/O2 ratio for balancing energy metabolism and carbon flux, which could be candidate targets for future metabolic engineering practice.

16.
Mater Sci Eng C Mater Biol Appl ; 109: 110615, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32228941

RESUMO

Four nanostructured MnO2 with various controllable morphologies, including nanowires, nanorods, nanotubes and nanoflowers were synthesized, and then further composited with nitrogen-doped graphene (NG) with the assistance of ultrasonication. The surface morphologies, phase structures, and electrochemical performances of the proposed MnO2/NG nanohybrids were investigated by various techniques, and their catalytic activities on the electrooxidation of dopamine (DA) and uric acid (UA) were compared systematically. The sensing performances were found to be highly correlated with their morphologies. Among these morphologies, the nanoflower-like MnO2, composited with NG, displayed the most sensitive response signals for DA and UA. The boosted electrocatalytic activity was ascribed to the unique porous structure, large electroactive area, and low charge transfer resistance (Rct), which facilitated the electron transfer between electrode and analytes. Two linear response ranges (0.1 µM-10 µM and 10 µM-100 µM) were accompanied with very low detection limits of 34 nM and 39 nM for DA and UA, respectively. Moreover, the successful application of the MnO2NFs/NG composites for the simultaneous detection of DA and UA in human serum was realized using second-derivative linear sweep voltammetry (SDLSV). These findings give valuable insights for understanding the morphology-dependent sensing properties of MnO2 based nanomaterials, which is conducive to the rapid development of ubiquitous MnO2-based electrochemical sensors.


Assuntos
Dopamina/sangue , Técnicas Eletroquímicas , Grafite/química , Compostos de Manganês/química , Nitrogênio/química , Óxidos/química , Ácido Úrico/sangue , Humanos
17.
Artigo em Inglês | MEDLINE | ID: mdl-32195227

RESUMO

Establishment of production platforms through prokaryotic engineering in microbial organisms would be one of the most efficient means for chemicals, protein, and biofuels production. Despite the fact that CRISPR (clustered regularly interspaced short palindromic repeats)-based technologies have readily emerged as powerful and versatile tools for genetic manipulations, their applications are generally limited in prokaryotes, possibly owing to the large size and severe cytotoxicity of the heterogeneous Cas (CRISPR-associated) effector. Nevertheless, the rich natural occurrence of CRISPR-Cas systems in many bacteria and most archaea holds great potential for endogenous CRISPR-based prokaryotic engineering. The endogenous CRISPR-Cas systems, with type I systems that constitute the most abundant and diverse group, would be repurposed as genetic manipulation tools once they are identified and characterized as functional in their native hosts. This article reviews the major progress made in understanding the mechanisms of invading DNA immunity by type I CRISPR-Cas and summarizes the practical applications of endogenous type I CRISPR-based toolkits for prokaryotic engineering.

18.
Artigo em Inglês | MEDLINE | ID: mdl-32195236

RESUMO

sRNAs represent a powerful class of regulators that influences multiple mRNA targets in response to environmental changes. However, very few direct sRNA-sRNA interactions have been deeply studied in any organism. Zymomonas mobilis is a bacterium with unique ethanol-producing metabolic pathways in which multiple small RNAs (sRNAs) have recently been identified, some of which show differential expression in ethanol stress. In this study, we show that two sRNAs (Zms4 and Zms6) are upregulated under ethanol stress and have significant impacts on ethanol tolerance and production in Z. mobilis. We conducted multi-omics analysis (combining transcriptomics and sRNA-immunoprecipitation) to map gene networks under the influence of their regulation. We confirmed that Zms4 and Zms6 bind multiple RNA targets and regulate their expressions, influencing many downstream pathways important to ethanol tolerance and production. In particular, Zms4 and Zms6 interact with each other as well as many other sRNAs, forming a novel sRNA-sRNA direct interaction network. This study thus uncovers a sRNA network that co-orchestrates multiple ethanol related pathways through a diverse set of mRNA targets and a large number of sRNAs. To our knowledge, this study represents one of the largest sRNA-sRNA direct interactions uncovered so far.

19.
Fitoterapia ; 142: 104531, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32114039

RESUMO

Nine new (1-9) and three known (10-12) sesquiterpenoids were isolated from the ethanol-water (7:3, v/v) extract of the Datura metel L. leaves. The structures of 1-9 were elucidated by detailed spectroscopic analyses, including 1D and 2D NMR, HR-ESI-MS. All isolates (1-12) were evaluated for anti-inflammatory activity against the production of nitrogen oxide in lipopolysaccharide-induced RAW264.7 cells and compound 5 possessed the best inhibitory effect among them, with the IC50 value reaching 9.33-11.67 µM, which was lower than positive control, L-NMMA, with IC50 range from 13.64 to 17.02 µM.


Assuntos
Anti-Inflamatórios/isolamento & purificação , Datura metel/química , Sesquiterpenos/isolamento & purificação , Animais , Anti-Inflamatórios/química , Camundongos , Células RAW 264.7 , Sesquiterpenos/química
20.
Front Microbiol ; 11: 13, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32038596

RESUMO

Zymomonas mobilis 8b is an ethanologenic bacterium engineered to utilize both glucose and xylose. The impacts of lignocellulosic hydrolyzate inhibitors on the growth of Zymomonas mobilis 8b have been investigated. However, the molecular responses of these inhibitors have not been completely elucidated yet. In this study, molecular responses to furfural were investigated using transcriptomic approaches of both chip-based microarray and a directional mRNA-Seq. Furfural acute shock time-course experiment with 3 g/L furfural supplemented when cells reached exponential phase and stress response experiment in the presence of 2 g/L furfural from the beginning of fermentation were carried out to study the physiological and transcriptional profiles of short-term and long-term effects of furfural on 8b. Furfural negatively affected 8b growth in terms of final biomass and the fermentation time. Transcriptomic studies indicated that the response of 8b to furfural was dynamic and complex, and differences existed between short-term shock and long-term stress responses. However, the gene function categories were similar with most down-regulated genes related to translation and biosynthesis, while the furfural up-regulated genes were mostly related to general stress responses. Several gene candidates have been identified and genetic studies indicated that expression of ZMO0465 and cysteine synthase operon ZMO0003-0006 driven by its native promoter in a shuttle vector enhanced the furfural tolerance of 8b. In addition, the relationship between microarray and mRNA-Seq was compared with good correlations. The directional mRNA-Seq data not only provided the gene expression profiling, but also can be applied for transcriptional architecture improvement to identify and confirm operons, novel transcripts, hypothetical gene functions, transcriptional start sites, and promoters with different strength.

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