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1.
Epidemiol Mikrobiol Imunol ; 68(2): 99-102, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31398983

RESUMO

The increasing incidence of multiresistant bacterial strains is currently a serious health concern. These pathogens are often the cause of nosocomial infections with limited treatment options and high fatality rates. A case report is presented of an uncommon detection of four different species (Citrobacter freundii, Klebsiella pneumoniae, Escherichia coli, and Morganella morganii) producing the same type of carbapenemase, KPC-2, in a female patient during her complicated long-term hospital stay. Resistance was probably spread to other species by horizontal transmission of plasmids carrying the blaKPC-2 genes. The implementation of strict anti-epidemic measures prevented further spread of these carbapenem-resistant bacteria.


Assuntos
Antibacterianos , Bactérias , Infecções Bacterianas , Infecção Hospitalar , beta-Lactamases , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/enzimologia , Infecções Bacterianas/microbiologia , Proteínas de Bactérias/genética , Coinfecção/microbiologia , Infecção Hospitalar/microbiologia , Farmacorresistência Bacteriana/genética , Feminino , Transferência Genética Horizontal , Humanos , Testes de Sensibilidade Microbiana , Plasmídeos/genética , beta-Lactamases/genética
2.
Yi Chuan ; 41(8): 736-745, 2019 Aug 20.
Artigo em Chinês | MEDLINE | ID: mdl-31447424

RESUMO

As one of plant cell wall components, pectin is the main anti-nutritional factor in livestock and poultry feeds and has an adverse effect on utilization efficiency of feed energy and nitrogen. Pectinases, which are widely found in microorganisms such as bacteria, yeast and filamentous fungi in nature,can improve feed efficiency by relieving the anti-nutritional effect of pectin through promoting the hydrolysis reaction of feed pectin. To explore the feasibility of expressing microbial-derived pectinase genes in pig cells, we introduced microbial-derived pectinase genes pg5a, pgI, pga3A, and pgaA into porcine PK 15 cells by lipofection for heterogenous expression. Enzymatic activities of the pectinases encoded by these genes were analyzed using the 3,5 dinitrosalicylic acid (DNS) method. Results showed that all four pectinase genes were able to be transcribed into mRNAs in porcine PK 15 cells, but only pg5a and pgI were adapted to the porcine cell expression system. Among them, the maximum activity of pectinase PG5A was 0.95 U/mL, the optimum pH was pH 4.0, and the enzymatic activity was maintained above 46% in the range of pH 4.6 to 6.0. Pectinase PGI obtained the highest enzymatic activity at pH 5.0, which was 0.30 U/mL, and maintained more than 35% of the activity in the range of pH 4.0 to 6.0. The results of digestive protease tolerance test showed that PG5A and PGI were highly resistant to pepsin and trypsin. After treatment with 1 mg/mL pig pepsin for two hours, the residual enzymatic activities of PG5A and PGI were 76% and 71%, respectively. And after two hours treatment with 1 mg/mL of pig trypsin, the remaining enzymatic activities of PG5A and PGI were 44% and 93%, respectively. In summary, pectinase PG5A and PGI can be effectively expressed in pig cells, and have strong tolerance to pig intestinal pH environment and digestive proteases. Therefore, both pg5a and pgI can be used as candidate genes for production of transgenic pigs.


Assuntos
Bactérias/enzimologia , Fungos/enzimologia , Poligalacturonase/biossíntese , Animais , Células Cultivadas , Pectinas , Poligalacturonase/genética , Suínos
3.
Nat Commun ; 10(1): 2917, 2019 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-31266949

RESUMO

Novel antibacterial agents are needed to address the emergence of global antibiotic resistance. MraY is a promising candidate for antibiotic development because it is the target of five classes of naturally occurring nucleoside inhibitors with potent antibacterial activity. Although these natural products share a common uridine moiety, their core structures vary substantially and they exhibit different activity profiles. An incomplete understanding of the structural and mechanistic basis of MraY inhibition has hindered the translation of these compounds to the clinic. Here we present crystal structures of MraY in complex with representative members of the liposidomycin/caprazamycin, capuramycin, and mureidomycin classes of nucleoside inhibitors. Our structures reveal cryptic druggable hot spots in the shallow inhibitor binding site of MraY that were not previously appreciated. Structural analyses of nucleoside inhibitor binding provide insights into the chemical logic of MraY inhibition, which can guide novel approaches to MraY-targeted antibiotic design.


Assuntos
Antibacterianos/química , Bactérias/enzimologia , Proteínas de Bactérias/química , Produtos Biológicos/química , Inibidores Enzimáticos/química , Nucleosídeos/antagonistas & inibidores , Transferases/química , Aminoglicosídeos/química , Arginina/análogos & derivados , Arginina/química , Bactérias/química , Bactérias/genética , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Cristalografia por Raios X , Transferases/antagonistas & inibidores , Transferases/genética , Transferases/metabolismo
4.
J Agric Food Chem ; 67(29): 8177-8185, 2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31290662

RESUMO

Trehalose synthase (TreS) catalyzes the reversible interconversion of maltose to trehalose, and is therefore essential for trehalose production. Consequently, dissecting the catalytic mechanism of TreS is important for enzyme optimization and industrial applications. TreS from Thermobaculum terrenum (TtTreS) is a thermostable enzyme. Here, we studied the composition of the TtTreS active site through computer calculation and enzyme analysis. The results were consistent with a two-step double-displacement mechanism, similar to that of glycoside hydrolase 13 family enzymes. However, our data suggested that glucose rotation, following breakage of the α-1,4 glycosidic bond, is a key factor determining the reaction direction and conversion rate. The N246 residue plays an important role in glucose rotation. Moreover, we established a saturated mutation model for the nonconserved amino acids around the substrate gateway domain. Finally, four TtTreS mutants (K136T, Y137D, K138N, and D139S) resulted in improved trehalose yield compared to that of the wild-type enzyme.


Assuntos
Bactérias/enzimologia , Proteínas de Bactérias/química , Glucosiltransferases/química , Bactérias/química , Bactérias/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Catálise , Domínio Catalítico , Biologia Computacional , Estabilidade Enzimática , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Temperatura Alta , Especificidade por Substrato
5.
Dokl Biochem Biophys ; 485(1): 107-110, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31201626

RESUMO

In this study, we formulated the principles of designing bioluminescent enzyme tests for assessing the quality of complex media, which consist in providing the maximum sensitivity to potentially toxic chemicals at a minimal impact of uncontaminated complex media. The developed principles served as a basis for designing a new bioluminescent method for an integrated rapid assessment of chemical safety of fruits and vegetables, which is based on using the luminous bacteria enzymes (NAD(P)H:FMN oxidoreductase and luciferase) as a test system.


Assuntos
Bactérias/enzimologia , Proteínas de Bactérias/química , FMN Redutase/química , Análise de Alimentos/métodos , Luciferases/química , Medições Luminescentes/métodos
6.
Future Microbiol ; 14: 671-689, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31161792

RESUMO

Aim: To propose newer combinations of antibiotics effective against NDM-1-producing bacterial strains. Materials & methods: Antibiotics combinations were tested by checkerboard assay. NDM-1 protein/enzyme was expressed and purified to perform enzyme kinetics, circular dichroism and fluorescence spectroscopic studies. Results: Doripenem-cefoxitin combination and doripenem-tetracycline combination showed synergistic effect toward NDM-1-producing strains. The catalytic efficiency of NDM-1 enzyme was decreased drastically by 96.6% upon doripenem-cefoxitin treatment and by 35.54% after doripenem-tetracycline treatment. Conformational changes were observed in NDM-1 upon combination treatment. Conclusion: NDM-1-producing bacterial strains show resistance to multiple antibiotics but the combination of doripenem-cefoxitin and doripenem-tetracycline are effective against them. The combination of a carbapenem and cephamycin antibiotic is proposed for future treatment options against bacteria-producing NDM-1.


Assuntos
Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Cefoxitina/farmacologia , Doripenem/farmacologia , Tetraciclina/farmacologia , beta-Lactamases/efeitos dos fármacos , Bactérias/enzimologia , Bactérias/metabolismo , Combinação de Medicamentos , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Sinergismo Farmacológico , Ensaios Enzimáticos , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Regulação Bacteriana da Expressão Gênica/genética , Cinética , Testes de Sensibilidade Microbiana , Inibidores da Síntese de Proteínas/farmacologia , Termodinâmica , beta-Lactamases/análise
7.
Subcell Biochem ; 92: 187-219, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31214988

RESUMO

Signal peptidases are the membrane bound enzymes that cleave off the amino-terminal signal peptide from secretory preproteins . There are two types of bacterial signal peptidases . Type I signal peptidase utilizes a serine/lysine catalytic dyad mechanism and is the major signal peptidase in most bacteria. Type II signal peptidase is an aspartic protease specific for prolipoproteins. This chapter will review what is known about the structure, function and mechanism of these unique enzymes.


Assuntos
Ácido Aspártico Endopeptidases/química , Ácido Aspártico Endopeptidases/metabolismo , Bactérias/enzimologia , Proteínas de Bactérias/química , Proteínas de Bactérias/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Serina Endopeptidases/química , Serina Endopeptidases/metabolismo
8.
Subcell Biochem ; 92: 223-274, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31214989

RESUMO

The Bacterial Phosphoenolpyruvate (PEP) : Sugar Phosphotransferase System (PTS) mediates the uptake and phosphorylation of carbohydrates, and controls the carbon- and nitrogen metabolism in response to the availability of sugars. PTS occur in eubacteria and in a few archaebacteria but not in animals and plants. All PTS comprise two cytoplasmic phosphotransferase proteins (EI and HPr) and a species-dependent, variable number of sugar-specific enzyme II complexes (IIA, IIB, IIC, IID). EI and HPr transfer phosphorylgroups from PEP to the IIA units. Cytoplasmic IIA and IIB units sequentially transfer phosphates to the sugar, which is transported by the IIC and IICIID integral membrane protein complexes. Phosphorylation by IIB and translocation by IIC(IID) are tightly coupled. The IIC(IID) sugar transporters of the PTS are in the focus of this review. There are four structurally different PTS transporter superfamilies (glucose, glucitol, ascorbate, mannose) . Crystal structures are available for transporters of two superfamilies: bcIICmal (MalT, 5IWS, 6BVG) and bcIICchb (ChbC, 3QNQ) of B. subtilis from the glucose family, and IICasc (UlaA, 4RP9, 5ZOV) of E. coli from the ascorbate superfamily . They are homodimers and each protomer has an independent transport pathway which functions by an elevator-type alternating-access mechanism. bcIICmal and bcIICchb have the same fold, IICasc has a completely different fold. Biochemical and biophysical data accumulated in the past with the transporters for mannitol (IICBAmtl) and glucose (IICBglc) are reviewed and discussed in the context of the bcIICmal crystal structures. The transporters of the mannose superfamily are dimers of protomers consisting of a IIC and a IID protein chain. The crystal structure is not known and the topology difficult to predict. Biochemical data indicate that the IICIID complex employs a different transport mechanism . Species specific IICIID serve as a gateway for the penetration of bacteriophage lambda DNA across, and insertion of class IIa bacteriocins into the inner membrane. PTS transporters are inserted into the membrane by SecYEG translocon and have specific lipid requirements. Immunoelectron- and fluorescence microscopy indicate a non-random distribution and supramolecular complexes of PTS proteins.


Assuntos
Bactérias/enzimologia , Bactérias/metabolismo , Sistema Fosfotransferase de Açúcar do Fosfoenolpiruvato/metabolismo , Açúcares/metabolismo , Transporte Biológico , Fosforilação
9.
Subcell Biochem ; 92: 301-335, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31214991

RESUMO

The invention of a biological membrane which is used as energy storage system to drive the metabolism of a primordial, unicellular organism represents a key event in the evolution of life. The innovative, underlying principle of this key event is respiration. In respiration, a lipid bilayer with insulating properties is chosen as the site for catalysis of an exergonic redox reaction converting substrates offered from the environment, using the liberated Gibbs free energy (ΔG) for the build-up of an electrochemical H+ (proton motive force, PMF) or Na+ gradient (sodium motive force, SMF) across the lipid bilayer. Very frequently , several redox reactions are performed in a consecutive manner, with the first reaction delivering a product which is used as substrate for the second redox reaction, resulting in a respiratory chain. From today's perspective, the (mostly) unicellular bacteria and archaea seem to be much simpler and less evolved when compared to multicellular eukaryotes. However, they are overwhelmingly complex with regard to the various respiratory chains which permit survival in very different habitats of our planet, utilizing a plethora of substances to drive metabolism. This includes nitrogen, sulfur and carbon compounds which are oxidized or reduced by specialized, respiratory enzymes of bacteria and archaea which lie at the heart of the geochemical N, S and C-cycles. This chapter gives an overview of general principles of microbial respiration considering thermodynamic aspects, chemical reactions and kinetic restraints. The respiratory chains of Escherichia coli and Vibrio cholerae are discussed as models for PMF- versus SMF-generating processes, respectively. We introduce main redox cofactors of microbial respiratory enzymes, and the concept of intra-and interelectron transfer. Since oxygen is an electron acceptor used by many respiratory chains, the formation and removal of toxic oxygen radicals is described. Promising directions of future research are respiratory enzymes as novel bacterial targets, and biotechnological applications relying on respiratory complexes.


Assuntos
Archaea/metabolismo , Bactérias/metabolismo , Membrana Celular/metabolismo , Transporte de Elétrons , Metabolismo Energético , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Archaea/citologia , Archaea/enzimologia , Bactérias/citologia , Bactérias/enzimologia
10.
Zhejiang Da Xue Xue Bao Yi Xue Ban ; 48(1): 44-49, 2019 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-31102357

RESUMO

Rifamycins, a group of bacterial RNA polymerase inhibitors, are the firstline antimicrobial drugs to treat tuberculosis. In light of the emergence of rifamycinresistant bacteria, development of new RNA polymerase inhibitors that kill rifamycinresistant bacteria with high bioavailability is urgent. Structural analysis of bacterial RNA polymerase in complex with inhibitors by crystallography and cryo-EM indicates that RNA polymerase inhibitors function through five distinct molecular mechanisms:inhibition of the extension of short RNA; competition with substrates; inhibition of the conformational change of the'bridge helix'; inhibition of clamp opening;inhibition of clamp closure. This article reviews the research progress of these five groups of RNA polymerase inhibitors to provide references for the modification of existing RNA polymerase inhibitors and the discovery of new RNA polymerase inhibitors.


Assuntos
RNA Polimerases Dirigidas por DNA , Descoberta de Drogas , RNA Bacteriano , Antituberculosos/uso terapêutico , Bactérias/efeitos dos fármacos , Bactérias/enzimologia , RNA Polimerases Dirigidas por DNA/metabolismo , Descoberta de Drogas/tendências , Farmacorresistência Bacteriana , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Humanos , Tuberculose/tratamento farmacológico , Tuberculose/enzimologia
11.
Talanta ; 201: 237-244, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31122418

RESUMO

Drug-resistant bacteria is posing one of the greatest threats to human health. Carbapenemase-producing (CP) bacteria are a group of emerging highly drug-resistant bacteria which cause serious health problem in worldwide. Rapid and reliable detection of CP-bacteria is essential for point-of-care therapy and rapid infection control. In this study, a high-throughput tip-desorption electrospray ionization (tip-DESI) with solid-substrate tip was developed to couple ion mobility-tandem mass spectrometry (IM-MS/MS) for rapid screening of CP-bacteria from clinical samples. Raw bacteria spiked with indicators (i.e., carbapenems) was directly loaded on disposable substrate tip that connected with high voltage, and a desorption spray was applied for desorption and ionization of analytes. The substrate materials and desorption/ionization modes were optimized in this study. CP bacteria were ambiguously identified by monitoring of characteristic IM drift time and MS/MS spectra of hydrolyzed and decarboxylated carbapenems. We demonstrated this method for direct detection CP-bacteria in complex samples, showing excellent analytical performances including good tolerance to complex matrices, reducing interferences, high specificity, good repeatability, high sensitivity, and high analytical speed. Furthermore, this method was also applied for fast screening of CP-bacteria from different clinical bacteria, showing the potential applications for fast and reliable detection of antibiotic resistance in clinics.


Assuntos
Bactérias/isolamento & purificação , Técnicas de Tipagem Bacteriana/métodos , Carbapenêmicos/urina , Espectrometria de Massas por Ionização por Electrospray/métodos , Bactérias/enzimologia , Proteínas de Bactérias/química , Carbapenêmicos/química , Resistência Microbiana a Medicamentos , Humanos , Hidrólise , Espectrometria de Mobilidade Iônica/métodos , Espectrometria de Massas em Tandem/métodos , beta-Lactamases/química
12.
mSphere ; 4(3)2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-31092601

RESUMO

Termites forage on a range of substrates, and it has been suggested that diet shapes the composition and function of termite gut bacterial communities. Through comparative analyses of gut metagenomes in nine termite species with distinct diets, we characterize bacterial community compositions and use peptide-based functional annotation method to determine biomass-degrading enzymes and the bacterial taxa that encode them. We find that fungus-growing termite guts have relatively more fungal cell wall-degrading enzyme genes, while wood-feeding termite gut communities have relatively more plant cell wall-degrading enzyme genes. Interestingly, wood-feeding termite gut bacterial genes code for abundant chitinolytic enzymes, suggesting that fungal biomass within the decaying wood likely contributes to gut bacterial or termite host nutrition. Across diets, the dominant biomass-degrading enzymes are predominantly coded for by the most abundant bacterial taxa, suggesting tight links between diet and gut community composition, with the most marked difference being the communities coding for the mycolytic capacity of the fungus-growing termite gut.IMPORTANCE Understanding functional capacities of gut microbiomes is important to improve our understanding of symbiotic associations. Here, we use peptide-based functional annotation to show that the gut microbiomes of fungus-farming termites code for a wealth of enzymes that likely target the fungal diet the termites eat. Comparisons to other termites showed that fungus-growing termite guts have relatively more fungal cell wall-degrading enzyme genes, whereas wood-feeding termite gut communities have relatively more plant cell wall-degrading enzyme genes. Across termites with different diets, the dominant biomass-degrading enzymes are predominantly coded for by the most abundant bacterial taxa, suggesting tight links between diet and gut community compositions.


Assuntos
Dieta , Fungos/crescimento & desenvolvimento , Microbioma Gastrointestinal , Isópteros/enzimologia , Isópteros/microbiologia , Animais , Bactérias/enzimologia , Bactérias/genética , Trato Gastrointestinal/enzimologia , Trato Gastrointestinal/microbiologia , Metagenoma , Filogenia , Células Vegetais/metabolismo , Análise de Sequência de DNA , Simbiose , Madeira/metabolismo
13.
Adv Exp Med Biol ; 1142: 115-129, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31102244

RESUMO

Lytic polysaccharide monooxygenases (LPMOs) are copper-dependent enzymes that catalyze the cleavage of 1,4-glycosidic bonds various plant cell wall polysaccharides and chitin. In contrast to glycoside hydrolases, LPMOs are active on the crystalline regions of polysaccharides and thus synergize with hydrolytic enzymes. This synergism leads to an overall increase in the biomass-degradation activity of enzyme mixtures. Chitin-active LPMOs were discovered in 2010 and are currently classified in families AA10, AA11, and AA15 of the Carbohydrate-Active enZYmes database, which include LPMOs from bacteria, fungi, insects, and viruses. LPMOs have become important enzymes both industrially and scientifically and, in this chapter, we provide a brief introduction to chitin-active LPMOs including a summary of the 20+ chitin-active LPMOs that have been characterized so far. Then, we describe their structural features, catalytic mechanism, and appended carbohydrate modules. Finally, we show how chitin-active LPMOs can be used to perform chemo-enzymatic modification of chitin substrates.


Assuntos
Quitina/química , Oxigenases de Função Mista , Animais , Bactérias/enzimologia , Parede Celular , Fungos/enzimologia , Glicosídeo Hidrolases , Insetos/enzimologia , Vírus/enzimologia
14.
Adv Exp Med Biol ; 1142: 131-151, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31102245

RESUMO

Chitin, a structural polysaccharide of ß-1,4-linked N-acetyl-D-glucosamine residues, is the second most abundant natural biopolymer after cellulose. The metabolism of chitin affects the global carbon and nitrogen cycles, which are maintained by marine and soil-dwelling bacteria. The degradation products of chitin metabolism serve as important nutrient sources for the chitinolytic bacteria. Chitinolytic bacteria have elaborate enzymatic systems for the degradation of the recalcitrant chitin biopolymer. This chapter introduces chitin degradation and utilization systems of the chitinolytic bacteria. These bacteria secrete many chitin-degrading enzymes, including processive chitinases, endo-acting non-processive chitinases, lytic polysaccharide monooxygenases, and N-acetyl-hexosaminidases. Bacterial chitinases play a fundamental role in the degradation of chitin. Enzymatic properties, catalytic mechanisms, and three-dimensional structures of chitinases have been extensively studied by many scientists. These enzymes can be exploited to produce a range of chitin-derived products, e.g., biocontrol agents against many plant pathogenic fungi and insects. We introduce bacterial chitinases in terms of their reaction modes and structural features.


Assuntos
Bactérias/enzimologia , Quitina/metabolismo , Quitinases/metabolismo , Oxigenases de Função Mista
15.
Bioelectrochemistry ; 129: 1-9, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31063949

RESUMO

This review summarizes the bioelectrocatalytic properties of d-fructose dehydrogenase (FDH), while taking into consideration its enzymatic characteristics. FDH is a membrane-bound flavohemo-protein with a molecular mass of 138 kDa, and it catalyzes the oxidation of d-fructose to 5-keto-d-fructose. The characteristic feature of FDH is its strong direct-electron-transfer (DET)-type bioelectrocatalytic activity. The pathway of the DET-type reaction is discussed. An overview of the application of FDH-based bioelectrocatalysis to biosensors and biofuel cells is also presented, and the benefits and problems associated with it are extensively discussed.


Assuntos
Bactérias/enzimologia , Fontes de Energia Bioelétrica , Técnicas Biossensoriais/métodos , Desidrogenases de Carboidrato/metabolismo , Frutose/metabolismo , Sequência de Aminoácidos , Bactérias/química , Bactérias/metabolismo , Biocatálise , Fontes de Energia Bioelétrica/microbiologia , Técnicas Biossensoriais/instrumentação , Desidrogenases de Carboidrato/química , Transporte de Elétrons , Oxirredução
16.
Sci Total Environ ; 678: 486-498, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31077927

RESUMO

Organic pollutants are continuously being introduced in seawater with uncharacterized impacts on the engines of the marine biogeochemical cycles, the microorganisms. The effects on marine microbial communities were assessed for perfluoroalkyl substances, organophosphate esters flame retardants and plasticizers, polycyclic aromatic hydrocarbons, and n-alkanes. Dose-response experiments were performed at three stations and at three depths in the NW Mediterranean with contrasted nutrient and pollutant concentrations. In these experiments, the microbial growth rates, the abundances of the main bacterial groups, measured by Catalyzed Reporter Deposition Fluorescence In Situ Hybridization (CARD-FISH), and extracellular enzymatic activities, were quantified. Increasing concentrations of organic pollutants (OPs) promoted different responses in the communities that were compound, organism and nutrient availability (trophic status). The largest differences between OP treatments and controls in the growth rates of both heterotrophic and phototrophic microbial groups were observed in seawater from the deep chlorophyll maxima. Furthermore, there was a compound specific stimulation of different extracellular enzymatic activities after the exposure to OPs. Our results revealed that marine microbial communities reacted not only to hydrocarbons, known to be used as a carbon source, but also to low concentrations of organic pollutants of emerging concern in a complex manner, reflecting the variability of various environmental variables. Multiple linear regressions suggested that organic pollutants modulated the bacterial growth and extracellular enzymatic activities, but this modulation was of lower magnitude than the observed pronounced response of the microbial community to nutrient availability.


Assuntos
Bactérias/efeitos dos fármacos , Hidrocarbonetos/efeitos adversos , Microbiota/efeitos dos fármacos , Poluentes Químicos da Água/efeitos adversos , Bactérias/enzimologia , Bactérias/crescimento & desenvolvimento , Mar Mediterrâneo , Compostos Orgânicos/efeitos adversos , Água do Mar/química , Espanha
17.
Int J Mol Sci ; 20(9)2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-31067645

RESUMO

The caseinolytic protease proteolytic subunit (ClpP) is a serine protease playing an important role in proteostasis of eukaryotic organelles and prokaryotic cells. Alteration of ClpP function has been proved to affect the virulence and infectivity of a number of pathogens. Increased bacterial resistance to antibiotics has become a global problem and new classes of antibiotics with novel mechanisms of action are needed. In this regard, ClpP has emerged as an attractive and potentially viable option to tackle pathogen fitness without suffering cross-resistance to established antibiotic classes and, when not an essential target, without causing an evolutionary selection pressure. This opens a greater window of opportunity for the host immune system to clear the infection by itself or by co-administration with commonly prescribed antibiotics. A comprehensive overview of the function, regulation and structure of ClpP across the different organisms is given. Discussion about mechanism of action of this protease in bacterial pathogenesis and human diseases are outlined, focusing on the compounds developed in order to target the activation or inhibition of ClpP.


Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Depsipeptídeos/farmacologia , Endopeptidase Clp/metabolismo , Antibacterianos/química , Bactérias/efeitos dos fármacos , Bactérias/enzimologia , Proteínas de Bactérias/agonistas , Proteínas de Bactérias/química , Depsipeptídeos/química , Desenho de Drogas , Endopeptidase Clp/química
18.
Appl Microbiol Biotechnol ; 103(12): 4679-4692, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31049619

RESUMO

Commercially, nitrilases are valuable biocatalysts capable of converting a diverse range of nitriles to carboxylic acids for the greener synthesis of chemicals and pharmaceuticals. Nitrilases are widespread in nature and are both important components of metabolic pathways and a response to environmental factors such as natural or manmade nitriles. Nitrilases are often grouped together on a genome in specific gene clusters that reflect these metabolic functions. Although nitrilase induction systems are still poorly understood, it is known that a powerful Rhodococcal transcription regulator system permits accumulation of intracellular nitrilase of up to 30-40% of total soluble protein in wild type Rhodococcous rhodochrous and host Streptomyces strains. Nitrilase expression inducer molecules encompass a broad range of aliphatic, aromatic and heteroaromatic nitriles, as well as some secondary and tertiary amides that are resistant to nitrilase degradation.


Assuntos
Aminoidrolases/biossíntese , Aminoidrolases/genética , Bactérias/enzimologia , Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Biocatálise , Indução Enzimática , Família Multigênica , Rhodococcus/enzimologia , Rhodococcus/genética , Streptomyces/enzimologia , Streptomyces/genética , Especificidade por Substrato
19.
Prep Biochem Biotechnol ; 49(8): 783-789, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31132010

RESUMO

To overcome the problem that soft-sensing model cannot be updated with the bioprocess changes, this article proposed a soft-sensing modeling method which combined fuzzy c-means clustering (FCM) algorithm with least squares support vector machine theory (LS-SVM). FCM is used for separating a whole training data set into several clusters with different centers, each subset is trained by LS-SVM and sub-models are developed to fit different hierarchical property of the process. The new sample data that bring new operation information is introduced in the model, and the fuzzy membership function of the sample to each clustering is first calculated by the FCM algorithm. Then, a corresponding LS-SVM sub-model of the clustering with the largest fuzzy membership function is used for performing dynamic learning so that the model can update online. The proposed method is applied to predict the key biological parameters in the marine alkaline protease MP process. The simulation result indicates that the soft-sensing modeling method increases the model's adaptive abilities in various operation conditions and can improve its generalization ability.


Assuntos
Organismos Aquáticos/enzimologia , Bactérias/enzimologia , Proteínas de Bactérias/metabolismo , Reatores Biológicos , Endopeptidases/metabolismo , Fermentação , Modelos Biológicos , Máquina de Vetores de Suporte , Algoritmos , Desenho de Equipamento , Análise dos Mínimos Quadrados
20.
Int J Mol Sci ; 20(9)2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31075847

RESUMO

A halo-thermophilic bacterium, Roseithermus sacchariphilus strain RA (previously known as Rhodothermaceae bacterium RA), was isolated from a hot spring in Langkawi, Malaysia. A complete genome analysis showed that the bacterium harbors 57 glycoside hydrolases (GHs), including a multi-domain xylanase (XynRA2). The full-length XynRA2 of 813 amino acids comprises a family 4_9 carbohydrate-binding module (CBM4_9), a family 10 glycoside hydrolase catalytic domain (GH10), and a C-terminal domain (CTD) for type IX secretion system (T9SS). This study aims to describe the biochemical properties of XynRA2 and the effects of CBM truncation on this xylanase. XynRA2 and its CBM-truncated variant (XynRA2ΔCBM) was expressed, purified, and characterized. The purified XynRA2 and XynRA2ΔCBM had an identical optimum temperature at 70 °C, but different optimum pHs of 8.5 and 6.0 respectively. Furthermore, XynRA2 retained 94% and 71% of activity at 4.0 M and 5.0 M NaCl respectively, whereas XynRA2ΔCBM showed a lower activity (79% and 54%). XynRA2 exhibited a turnover rate (kcat) of 24.8 s-1, but this was reduced by 40% for XynRA2ΔCBM. Both the xylanases hydrolyzed beechwood xylan predominantly into xylobiose, and oat-spelt xylan into a mixture of xylo-oligosaccharides (XOs). Collectively, this work suggested CBM4_9 of XynRA2 has a role in enzyme performance.


Assuntos
Bactérias/enzimologia , Endo-1,4-beta-Xilanases/química , Endo-1,4-beta-Xilanases/metabolismo , Variação Genética , Proteínas Mutantes/metabolismo , Tolerância ao Sal , Sequência de Aminoácidos , Endo-1,4-beta-Xilanases/genética , Cinética , Proteínas Mutantes/química , Filogenia , Domínios Proteicos , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Eletricidade Estática , Especificidade por Substrato , Xilanos/metabolismo
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