Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 88.437
Filtrar
1.
Curr Microbiol ; 78(10): 3696-3707, 2021 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-34426858

RESUMEN

We previously reported that the two-component system MacRS regulates morphogenesis and production of the blue-pigmented antibiotic actinorhodin (ACT) in Streptomyces coelicolor. In this study, the role of MacRS was further extended to include control of the production of the red-pigmented antibiotic undecylprodigiosin (RED) and the calcium-dependent antibiotic (CDA), and control of other important cellular activities. Our data indicated that disruption of the MacRS TCS reduced production not only of ACT but also of RED and CDA. RNA-Seq analysis revealed that genes involved in both secondary metabolism and primary metabolism are differentially expressed in the MacRS deletion mutant ΔmacRS. Moreover, we found that genes of the Zur regulon are also markedly downregulated in ΔmacRS, suggesting a role for macRS in zinc homeostasis. In addition to previously identified MacR sites with strong matches to the MacR consensus recognition sequence, a genome-wide search revealed over one hundred less-stringent matches, including potential sites upstream of absR1, crgA, and smeA. Electrophoretic mobility shift assays demonstrated that MacR binds some of these sites in vitro. Although there is no strong MacR site upstream of the ACT regulatory gene actII-orf4 (sco5085), we showed that an engineered MacR site enhanced ACT production, providing an approach for modulating production of useful compounds. Altogether, our work suggests an important role for MacRS in a range of cellular activities in Streptomyces and its potential application in strain engineering.


Asunto(s)
Streptomyces coelicolor , Antraquinonas , Antibacterianos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Regulación Bacteriana de la Expresión Génica , Regulón , Streptomyces coelicolor/genética , Streptomyces coelicolor/metabolismo
2.
Int J Mol Sci ; 22(16)2021 Aug 19.
Artículo en Inglés | MEDLINE | ID: mdl-34445643

RESUMEN

The development and spread of antibiotics and biocides resistance is a significant global challenge. To find a solution for this emerging problem, the discovery of novel bacterial cellular targets and the critical pathways associated with antimicrobial resistance is needed. In the present study, we investigated the role of the two most critical envelope stress response regulators, RpoE and CpxR, on the physiology and susceptibility of growing Salmonella enterica serovar enteritidis cells using the polycationic antimicrobial agent, chlorhexidine (CHX). It was shown that deletion of the cpxR gene significantly increased the susceptibility of this organism, whereas deletion of the rpoE gene had no effect on the pathogen's susceptibility to this antiseptic. It has been shown that a lack of the CpxR regulator induces multifaceted stress responses not only in the envelope but also in the cytosol, further affecting the key biomolecules, including DNA, RNA, and proteins. We showed that alterations in cellular trafficking and most of the stress responses are associated with a dysfunctional CpxR regulator during exponential growth phase, indicating that these physiological changes are intrinsically associated with the lack of the CpxR regulator. In contrast, induction of type II toxin-antitoxin systems and decrease of abundances of enzymes and proteins associated with the recycling of muropeptides and resistance to polymixin and cationic antimicrobial peptides were specific responses of the ∆cpxR mutant to the CHX treatment. Overall, our study provides insight into the effects of CpxR on the physiology of S. Enteritidis cells during the exponential growth phase and CHX treatment, which may point to potential cellular targets for the development of an effective antimicrobial agent.


Asunto(s)
Antiinfecciosos Locales/farmacología , Proteínas Bacterianas/metabolismo , Clorhexidina/farmacología , Regulación Bacteriana de la Expresión Génica , Salmonella enteritidis/crecimiento & desarrollo , Proteínas Bacterianas/genética , Mutación , Proteoma/análisis , Proteoma/metabolismo , Salmonella enteritidis/efectos de los fármacos , Salmonella enteritidis/metabolismo
3.
Microb Pathog ; 159: 105119, 2021 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-34339796

RESUMEN

Staphylococcus aureus is an eminent and opportunistic human pathogen that can colonize in the intestines, skin tissue and perineal regions of the host and cause severe infectious diseases. The presence of complex regulatory network and existence of virulent gene expression along with tuning metabolism enables the S. aureus to adopt the diversity of environments. Two component system (TCS) is a widely distributed mechanism in S. aureus that permit it for changing gene expression profile in response of environment stimuli. TCS usually consist of transmembrane histidine kinase (HK) and cytosolic response regulator. S. aureus contains totally 16 conserved pairs of two component systems, involving in different signaling mechanisms. There is a connection among these regulatory circuits and they can easily have effect on each other's expression. This review has discussed five major types of TCS in S. aureus and covers the recent knowledge of their virulence gene expression. We can get more understanding towards staphylococcal pathogenicity by getting insights about gene regulatory pathways via TCS, which can further provide implications in vaccine formation and new ways for drug design to combat serious infections caused by S. aureus in humans.


Asunto(s)
Infecciones Estafilocócicas , Staphylococcus aureus , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Regulación Bacteriana de la Expresión Génica , Histidina Quinasa/genética , Humanos , Staphylococcus aureus/genética , Staphylococcus aureus/metabolismo , Virulencia
4.
Molecules ; 26(15)2021 Jul 29.
Artículo en Inglés | MEDLINE | ID: mdl-34361735

RESUMEN

Biofilms, the predominant growth mode of microorganisms, pose a significant risk to human health. The protective biofilm matrix, typically composed of exopolysaccharides, proteins, nucleic acids, and lipids, combined with biofilm-grown bacteria's heterogenous physiology, leads to enhanced fitness and tolerance to traditional methods for treatment. There is a need to identify biofilm inhibitors using diverse approaches and targeting different stages of biofilm formation. This review discusses discovery strategies that successfully identified a wide range of inhibitors and the processes used to characterize their inhibition mechanism and further improvement. Additionally, we examine the structure-activity relationship (SAR) for some of these inhibitors to optimize inhibitor activity.


Asunto(s)
Antibacterianos/farmacología , Biopelículas/efectos de los fármacos , Matriz Extracelular de Sustancias Poliméricas/efectos de los fármacos , Bacterias Gramnegativas/efectos de los fármacos , Bacterias Grampositivas/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Antibacterianos/biosíntesis , Antibacterianos/síntesis química , Antibacterianos/aislamiento & purificación , Proteínas Bacterianas/antagonistas & inhibidores , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Biopelículas/crecimiento & desarrollo , GMP Cíclico/antagonistas & inhibidores , GMP Cíclico/química , GMP Cíclico/metabolismo , Diseño de Fármacos , Descubrimiento de Drogas , Farmacorresistencia Bacteriana/efectos de los fármacos , Matriz Extracelular de Sustancias Poliméricas/química , Matriz Extracelular de Sustancias Poliméricas/metabolismo , Bacterias Gramnegativas/crecimiento & desarrollo , Bacterias Gramnegativas/patogenicidad , Bacterias Grampositivas/crecimiento & desarrollo , Bacterias Grampositivas/patogenicidad , Lípidos/antagonistas & inhibidores , Lípidos/química , Pruebas de Sensibilidad Microbiana , Ácidos Nucleicos/antagonistas & inhibidores , Ácidos Nucleicos/química , Ácidos Nucleicos/metabolismo , Polisacáridos Bacterianos/antagonistas & inhibidores , Polisacáridos Bacterianos/química , Polisacáridos Bacterianos/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/aislamiento & purificación , Relación Estructura-Actividad
5.
Nat Commun ; 12(1): 4929, 2021 08 13.
Artículo en Inglés | MEDLINE | ID: mdl-34389727

RESUMEN

Synthetic metabolic pathways are a burden for engineered bacteria, but the underlying mechanisms often remain elusive. Here we show that the misregulated activity of the transcription factor Cra is responsible for the growth burden of glycerol overproducing E. coli. Glycerol production decreases the concentration of fructose-1,6-bisphoshate (FBP), which then activates Cra resulting in the downregulation of glycolytic enzymes and upregulation of gluconeogenesis enzymes. Because cells grow on glucose, the improper activation of gluconeogenesis and the concomitant inhibition of glycolysis likely impairs growth at higher induction of the glycerol pathway. We solve this misregulation by engineering a Cra-binding site in the promoter controlling the expression of the rate limiting enzyme of the glycerol pathway to maintain FBP levels sufficiently high. We show the broad applicability of this approach by engineering Cra-dependent regulation into a set of constitutive and inducible promoters, and use one of them to overproduce carotenoids in E. coli.


Asunto(s)
Escherichia coli/genética , Glucólisis/genética , Ingeniería Metabólica/métodos , Metabolómica/métodos , Proteómica/métodos , Transcripción Genética , Algoritmos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Sitios de Unión/genética , Carotenoides/metabolismo , Escherichia coli/metabolismo , Regulación Bacteriana de la Expresión Génica , Modelos Genéticos , Regiones Promotoras Genéticas/genética , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo
6.
Science ; 373(6556): 768-774, 2021 08 13.
Artículo en Inglés | MEDLINE | ID: mdl-34385391

RESUMEN

CRISPR-associated transposition systems allow guide RNA-directed integration of a single DNA cargo in one orientation at a fixed distance from a programmable target sequence. We used cryo-electron microscopy (cryo-EM) to define the mechanism that underlies this process by characterizing the transposition regulator, TnsC, from a type V-K CRISPR-transposase system. In this scenario, polymerization of adenosine triphosphate-bound TnsC helical filaments could explain how polarity information is passed to the transposase. TniQ caps the TnsC filament, representing a universal mechanism for target information transfer in Tn7/Tn7-like elements. Transposase-driven disassembly establishes delivery of the element only to unused protospacers. Finally, TnsC transitions to define the fixed point of insertion, as revealed by structures with the transition state mimic ADP•AlF3 These mechanistic findings provide the underpinnings for engineering CRISPR-associated transposition systems for research and therapeutic applications.


Asunto(s)
Proteínas Bacterianas/química , Proteínas Asociadas a CRISPR/química , Cianobacterias/química , Elementos Transponibles de ADN , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Proteínas Bacterianas/metabolismo , Proteínas Asociadas a CRISPR/metabolismo , Microscopía por Crioelectrón , Cianobacterias/genética , Cianobacterias/metabolismo , ADN Bacteriano/metabolismo , Modelos Moleculares , Conformación Proteica , Pliegue de Proteína , ARN Bacteriano/metabolismo , Transposasas/química , Transposasas/metabolismo
7.
Front Cell Infect Microbiol ; 11: 698146, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34368016

RESUMEN

L-arabinose inducible promoters are commonly used in gene expression analysis. However, nutrient source and availability also play a role in biofilm formation; therefore, L-arabinose metabolism could impact biofilm development. In this study we examined the impact of L-arabinose on Salmonella enterica serovar Typhimurium (S. Typhimurium) biofilm formation. Using mutants impaired for the transport and metabolism of L-arabinose, we showed that L-arabinose metabolism negatively impacts S. Typhimurium biofilm formation in vitro. When L-arabinose metabolism is abrogated, biofilm formation returned to baseline levels. However, without the ability to import extracellular L-arabinose, biofilm formation significantly increased. Using RNA-Seq we identified several gene families involved in these different phenotypes including curli expression, amino acid synthesis, and L-arabinose metabolism. Several individual candidate genes were tested for their involvement in the L-arabinose-mediated biofilm phenotypes, but most played no significant role. Interestingly, in the presence of L-arabinose the diguanylate cyclase gene adrA was downregulated in wild type S. Typhimurium. Meanwhile cyaA, encoding an adenylate cyclase, was downregulated in an L-arabinose transport mutant. Using an IPTG-inducible plasmid to deplete c-di-GMP via vieA expression, we were able to abolish the increased biofilm phenotype seen in the transport mutant. However, the mechanism by which the L-arabinose import mutant forms significantly larger biofilms remains to be determined. Regardless, these data suggest that L-arabinose metabolism influences intracellular c-di-GMP levels and therefore biofilm formation. These findings are important when considering the use of an L-arabinose inducible promoter in biofilm conditions.


Asunto(s)
Arabinosa , Proteínas Bacterianas , Biopelículas , Salmonella typhimurium , Arabinosa/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , GMP Cíclico , Regulación Bacteriana de la Expresión Génica , Plásmidos , Salmonella typhimurium/genética , Salmonella typhimurium/metabolismo
8.
Adv Protein Chem Struct Biol ; 127: 343-364, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34340773

RESUMEN

BACKGROUND AND AIM: The persistence of extensively drug-resistant (XDR) strains of Mycobacterium tuberculosis (MTB) continue to pose a significant challenge to the treatment and control of tuberculosis infections worldwide. XDR-MTB strains exhibit resistance against first-line anti-TB drugs, fluoroquinolones, and second-line injectable drugs. The mechanisms of drug resistance of MTB remains poorly understood. Our study aims at identifying the differentially expressed genes (DEGs), associated gene networks, and signaling cascades involved in rendering this pathogen resistant to multiple drugs, namely, isoniazid, rifampicin, and capreomycin. METHODS: We used the microarray dataset GSE53843. The GEO2R tool was used to prioritize the most significant DEGs (top 250) of each drug exposure sample between XDR strains and non-resistant strains. The validation of the 250 DEGs was performed using volcano plots. Protein-protein interaction networks of the DEGs were created using STRING and Cytoscape tools, which helped decipher the relationship between these genes. The significant DEGs were functionally annotated using DAVID and ClueGO. The concomitant biological processes (BP) and molecular functions (MF) were represented as dot plots. RESULTS AND CONCLUSION: We identified relevant molecular pathways and biological processes, such as cell wall biogenesis, lipid metabolic process, ion transport, phosphopantetheine binding, and triglyceride lipase activity. These processes indicated the involvement of multiple interconnected mechanisms in drug resistance. Our study highlighted the impact of cell wall permeability, with the dysregulation of the mur family of proteins, as essential factors in the inference of resistance. Additionally, upregulation of genes responsible for ion transport such as ctpF, arsC, and nark3, emphasizes the importance of transport channels and efflux pumps in potentially driving out stress-inducing compounds. This study investigated the upregulation of the Lip family of proteins, which play a crucial role in triglyceride lipase activity. Thereby illuminating the potential role of drug-induced dormancy and subsequent resistance in the mycobacterial strains. Multiple mechanisms such as carboxylic acid metabolic process, NAD biosynthetic process, triglyceride lipase activity, phosphopantetheine binding, organic acid biosynthetic process, and growth of symbiont in host cell were observed to partake in resistance of XDR-MTB. This study ultimately provides a platform for important mapping targets for potential therapeutics against XDR-MTB.


Asunto(s)
Proteínas Bacterianas , Farmacorresistencia Bacteriana/genética , Tuberculosis Extensivamente Resistente a Drogas , Regulación Bacteriana de la Expresión Génica , Mycobacterium tuberculosis , Biología de Sistemas , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Tuberculosis Extensivamente Resistente a Drogas/genética , Tuberculosis Extensivamente Resistente a Drogas/metabolismo , Humanos
9.
Nat Commun ; 12(1): 5065, 2021 08 20.
Artículo en Inglés | MEDLINE | ID: mdl-34417452

RESUMEN

The widespread UbiD enzyme family utilises the prFMN cofactor to achieve reversible decarboxylation of acrylic and (hetero)aromatic compounds. The reaction with acrylic compounds based on reversible 1,3-dipolar cycloaddition between substrate and prFMN occurs within the confines of the active site. In contrast, during aromatic acid decarboxylation, substantial rearrangement of the substrate aromatic moiety associated with covalent catalysis presents a molecular dynamic challenge. Here we determine the crystal structures of the multi-subunit vanillic acid decarboxylase VdcCD. We demonstrate that the small VdcD subunit acts as an allosteric activator of the UbiD-like VdcC. Comparison of distinct VdcCD structures reveals domain motion of the prFMN-binding domain directly affects active site architecture. Docking of substrate and prFMN-adduct species reveals active site reorganisation coupled to domain motion supports rearrangement of the substrate aromatic moiety. Together with kinetic solvent viscosity effects, this establishes prFMN covalent catalysis of aromatic (de)carboxylation is afforded by UbiD dynamics.


Asunto(s)
Carboxiliasas/química , Carboxiliasas/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Biocatálisis , Dominio Catalítico , Reacción de Cicloadición , Descarboxilación , Mononucleótido de Flavina/metabolismo , Cinética , Modelos Moleculares , Oxígeno/farmacología , Dominios Proteicos , Subunidades de Proteína/química , Subunidades de Proteína/metabolismo , Solventes , Relación Estructura-Actividad , Especificidad por Sustrato , Viscosidad
10.
Structure ; 29(8): 781-782, 2021 08 05.
Artículo en Inglés | MEDLINE | ID: mdl-34358463

RESUMEN

The bacterial Sec translocase transports unfolded proteins across membranes. In this issue of Structure, Krishnamurthy et al. (2021) report a nexus of conformational dynamics in the translocase motor protein, SecA. Their findings shed light on the Sec activation mechanism and suggest a general role for multi-level dynamics in protein functions.


Asunto(s)
Proteínas Bacterianas , Proteínas de Transporte de Membrana , Adenosina Trifosfatasas/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Transporte de Proteínas , Canales de Translocación SEC/genética , Proteína SecA
11.
Nat Commun ; 12(1): 4748, 2021 08 06.
Artículo en Inglés | MEDLINE | ID: mdl-34362927

RESUMEN

Encapsulins are a class of microbial protein compartments defined by the viral HK97-fold of their capsid protein, self-assembly into icosahedral shells, and dedicated cargo loading mechanism for sequestering specific enzymes. Encapsulins are often misannotated and traditional sequence-based searches yield many false positive hits in the form of phage capsids. Here, we develop an integrated search strategy to carry out a large-scale computational analysis of prokaryotic genomes with the goal of discovering an exhaustive and curated set of all HK97-fold encapsulin-like systems. We find over 6,000 encapsulin-like systems in 31 bacterial and four archaeal phyla, including two novel encapsulin families. We formulate hypotheses about their potential biological functions and biomedical relevance, which range from natural product biosynthesis and stress resistance to carbon metabolism and anaerobic hydrogen production. An evolutionary analysis of encapsulins and related HK97-type virus families shows that they share a common ancestor, and we conclude that encapsulins likely evolved from HK97-type bacteriophages.


Asunto(s)
Bacteriófagos/metabolismo , Cápside/metabolismo , Células Procariotas/metabolismo , Células Procariotas/virología , Archaea/metabolismo , Bacterias/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Bacteriófagos/genética , Evolución Biológica , Vías Biosintéticas/genética , Proteínas de la Cápside/química , Proteínas de la Cápside/metabolismo , Virus ADN/metabolismo , Filogenia , Virulencia
12.
J Phys Chem Lett ; 12(32): 7878-7884, 2021 Aug 19.
Artículo en Inglés | MEDLINE | ID: mdl-34382809

RESUMEN

The histone-like nucleoid structuring (H-NS) protein controls the expression of hundreds of genes in Gram-positive bacteria through its capability to coat and condense DNA. This mechanism requires the formation of superhelical H-NS protein filaments that are sensitive to temperature and salinity, allowing H-NS to act as an environment sensor. We use multiscale modeling and simulations to obtain detailed insights into the mechanism of H-NS filament's sensitivity to environmental changes. Through the simulations of the superhelical H-NS filament, we reveal how different environments induce heterogeneity of H-NS monomers. Further, we observe that transient self-association within the H-NS filament creates temperature-inducible strain and might mildly oppose DNA binding. We also probe different H-NS-DNA complex architectures and show that complexation enhances the stability of both DNA and H-NS superhelices. Overall, our results provide unprecedented molecular insights into the environmental sensing and DNA interactions of a prototypical nucleoid-structuring bacterial protein filament.


Asunto(s)
Proteínas Bacterianas/química , Proteínas de Unión al ADN/química , Secuencia de Aminoácidos , Proteínas Bacterianas/metabolismo , ADN/química , ADN/metabolismo , Proteínas de Unión al ADN/metabolismo , Simulación de Dinámica Molecular , Unión Proteica , Dominios Proteicos , Multimerización de Proteína , Salinidad , Salmonella typhimurium/química , Temperatura
13.
BMC Genomics ; 22(1): 633, 2021 Aug 31.
Artículo en Inglés | MEDLINE | ID: mdl-34461836

RESUMEN

BACKGROUND: Halogenation is a recurring feature in natural products, especially those from marine organisms. The selectivity with which halogenating enzymes act on their substrates renders halogenases interesting targets for biocatalyst development. Recently, CylC - the first predicted dimetal-carboxylate halogenase to be characterized - was shown to regio- and stereoselectively install a chlorine atom onto an unactivated carbon center during cylindrocyclophane biosynthesis. Homologs of CylC are also found in other characterized cyanobacterial secondary metabolite biosynthetic gene clusters. Due to its novelty in biological catalysis, selectivity and ability to perform C-H activation, this halogenase class is of considerable fundamental and applied interest. The study of CylC-like enzymes will provide insights into substrate scope, mechanism and catalytic partners, and will also enable engineering these biocatalysts for similar or additional C-H activating functions. Still, little is known regarding the diversity and distribution of these enzymes. RESULTS: In this study, we used both genome mining and PCR-based screening to explore the genetic diversity of CylC homologs and their distribution in bacteria. While we found non-cyanobacterial homologs of these enzymes to be rare, we identified a large number of genes encoding CylC-like enzymes in publicly available cyanobacterial genomes and in our in-house culture collection of cyanobacteria. Genes encoding CylC homologs are widely distributed throughout the cyanobacterial tree of life, within biosynthetic gene clusters of distinct architectures (combination of unique gene groups). These enzymes are found in a variety of biosynthetic contexts, which include fatty-acid activating enzymes, type I or type III polyketide synthases, dialkylresorcinol-generating enzymes, monooxygenases or Rieske proteins. Our study also reveals that dimetal-carboxylate halogenases are among the most abundant types of halogenating enzymes in the phylum Cyanobacteria. CONCLUSIONS: Our data show that dimetal-carboxylate halogenases are widely distributed throughout the Cyanobacteria phylum and that BGCs encoding CylC homologs are diverse and mostly uncharacterized. This work will help guide the search for new halogenating biocatalysts and natural product scaffolds.


Asunto(s)
Proteínas Bacterianas , Cianobacterias , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Cianobacterias/genética , Cianobacterias/metabolismo , Halogenación , Humanos , Familia de Multigenes , Recurrencia Local de Neoplasia
14.
Int J Mol Sci ; 22(15)2021 Jul 31.
Artículo en Inglés | MEDLINE | ID: mdl-34361010

RESUMEN

Biofilms are complex structures formed by a community of microbes adhering to a surface and/or to each other through the secretion of an adhesive and protective matrix. The establishment of these structures requires a coordination of action between microorganisms through powerful communication systems such as quorum-sensing. Therefore, auxiliary bacteria capable of interfering with these means of communication could be used to prevent biofilm formation and development. The phytopathogen Rhizobium rhizogenes, which causes hairy root disease and forms large biofilms in hydroponic crops, and the biocontrol agent Rhodococcus erythropolis R138 were used for this study. Changes in biofilm biovolume and structure, as well as interactions between rhizobia and rhodococci, were monitored by confocal laser scanning microscopy with appropriate fluorescent biosensors. We obtained direct visual evidence of an exchange of signals between rhizobia and the jamming of this communication by Rhodococcus within the biofilm. Signaling molecules were characterized as long chain (C14) N-acyl-homoserine lactones. The role of the Qsd quorum-quenching pathway in biofilm alteration was confirmed with an R. erythropolis mutant unable to produce the QsdA lactonase, and by expression of the qsdA gene in a heterologous host, Escherichia coli. Finally, Rhizobium biofilm formation was similarly inhibited by a purified extract of QsdA enzyme.


Asunto(s)
Agrobacterium/fisiología , Biopelículas , Percepción de Quorum , Rhodococcus/fisiología , Acil-Butirolactonas/metabolismo , Agrobacterium/genética , Agrobacterium/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Hidrolasas de Éster Carboxílico/genética , Hidrolasas de Éster Carboxílico/metabolismo , Rhodococcus/genética , Rhodococcus/metabolismo
15.
Int J Mol Sci ; 22(15)2021 Jul 30.
Artículo en Inglés | MEDLINE | ID: mdl-34360977

RESUMEN

Inhibition of ruminal microbial urease is of particular interest due to its crucial role in regulating urea-N utilization efficiency and nitrogen pollution in the livestock industry. Acetohydroxamic acid (AHA) is currently the only commercially available urease inhibitor, but it has adverse side effects. The urease accessory protein UreG, which facilitates the functional incorporation of the urease nickel metallocentre, has been proposed in developing urease inhibitor through disrupting urease maturation. The objective of this study was to screen natural compounds as potential urease inhibitors by targeting UreG in a predominant ruminal microbial urease. In silico screening and in vitro tests for potential inhibitors were performed using molecular docking and an assay for the GTPase activity of UreG. Chelerythrine chloride was selected as a potential urease inhibitor of UreG with an inhibition concentration IC50 value of 18.13 µM. It exhibited mixed inhibition, with the Ki value being 26.28 µM. We further explored its inhibition mechanism using isothermal titration calorimetry (ITC) and circular dichroism (CD) spectroscopy, and we found that chelerythrine chloride inhibited the binding of nickel to UreG and induced changes in the secondary structure, especially the α-helix and ß-sheet of UreG. Chelerythrine chloride formed a pi-anion interaction with the Asp41 residue of UreG, which is an important residue in initiating the conformational changes of UreG. In conclusion, chelerythrine chloride exhibited a potential inhibitory effect on urease, which provided new evidence for strategies to develop novel urease inhibitors targeting UreG to reduce nitrogen excretion from ruminants.


Asunto(s)
Proteínas Bacterianas/antagonistas & inhibidores , Benzofenantridinas/farmacología , Inhibidores Enzimáticos/farmacología , Proteínas de Unión a Fosfato/antagonistas & inhibidores , Rumen/microbiología , Amoníaco/metabolismo , Animales , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Benzofenantridinas/química , Sitios de Unión , Bovinos , Inhibidores Enzimáticos/química , Concentración 50 Inhibidora , Simulación del Acoplamiento Molecular , Proteínas de Unión a Fosfato/química , Proteínas de Unión a Fosfato/metabolismo , Unión Proteica
16.
Nat Commun ; 12(1): 4707, 2021 08 04.
Artículo en Inglés | MEDLINE | ID: mdl-34349110

RESUMEN

Salmonella utilizes translocated virulence proteins (termed effectors) to promote host cell invasion. The effector SopD contributes to invasion by promoting scission of the plasma membrane, generating Salmonella-containing vacuoles. SopD is expressed in all Salmonella lineages and plays important roles in animal models of infection, but its host cell targets are unknown. Here we show that SopD can bind to and inhibit the small GTPase Rab10, through a C-terminal GTPase activating protein (GAP) domain. During infection, Rab10 and its effectors MICAL-L1 and EHBP1 are recruited to invasion sites. By inhibiting Rab10, SopD promotes removal of Rab10 and recruitment of Dynamin-2 to drive scission of the plasma membrane. Together, our study uncovers an important role for Rab10 in regulating plasma membrane scission and identifies the mechanism used by a bacterial pathogen to manipulate this function during infection.


Asunto(s)
Proteínas Bacterianas/metabolismo , Membrana Celular/metabolismo , Salmonella typhimurium/patogenicidad , Proteínas de Unión al GTP rab/antagonistas & inhibidores , Proteínas Bacterianas/genética , Dinamina II , Proteínas Activadoras de GTPasa/genética , Proteínas Activadoras de GTPasa/metabolismo , Células HEK293 , Humanos , Salmonella typhimurium/metabolismo , Vacuolas/metabolismo , Vacuolas/microbiología , Virulencia , Proteínas de Unión al GTP rab/genética , Proteínas de Unión al GTP rab/metabolismo
17.
Int J Mol Sci ; 22(15)2021 Jul 22.
Artículo en Inglés | MEDLINE | ID: mdl-34360611

RESUMEN

Due to the high exposition to changing environmental conditions, bacteria have developed many mechanisms enabling immediate adjustments of gene expression. In many cases, the required speed and plasticity of the response are provided by RNA-dependent regulatory mechanisms. This is possible due to the very high dynamics and flexibility of an RNA structure, which provide the necessary sensitivity and specificity for efficient sensing and transduction of environmental signals. In this review, we will discuss the current knowledge about known bacterial regulatory mechanisms which rely on RNA structure. To better understand the structure-driven modulation of gene expression, we describe the basic theory on RNA structure folding and dynamics. Next, we present examples of multiple mechanisms employed by RNA regulators in the control of bacterial transcription and translation.


Asunto(s)
Bacterias/genética , Proteínas Bacterianas/metabolismo , Regulación Bacteriana de la Expresión Génica , Conformación de Ácido Nucleico , Pliegue del ARN , ARN Bacteriano/química , Bacterias/crecimiento & desarrollo , Bacterias/metabolismo , Proteínas Bacterianas/genética , Transcripción Genética
18.
Int J Mol Sci ; 22(15)2021 Jul 22.
Artículo en Inglés | MEDLINE | ID: mdl-34360615

RESUMEN

In contrast to Bacillus subtilis, Streptomyces coelicolor A3(2) contains nine homologues of stress response sigma factor SigB with a major role in differentiation and osmotic stress response. The aim of this study was to further characterize these SigB homologues. We previously established a two-plasmid system to identify promoters recognized by sigma factors and used it to identify promoters recognized by the three SigB homologues, SigF, SigG, and SigH from S. coelicolor A3(2). Here, we used this system to identify 14 promoters recognized by SigB. The promoters were verified in vivo in S. coelicolor A3(2) under osmotic stress conditions in sigB and sigH operon mutants, indicating some cross-recognition of these promoters by these two SigB homologues. This two-plasmid system was used to examine the recognition of all identified SigB-, SigF-, SigG-, and SigH-dependent promoters with all nine SigB homologues. The results confirmed this cross-recognition. Almost all 24 investigated promoters were recognized by two or more SigB homologues and data suggested some distinguishing groups of promoters recognized by these sigma factors. However, analysis of the promoters did not reveal any specific sequence characteristics for these recognition groups. All promoters showed high similarity in the -35 and -10 regions. Immunoblot analysis revealed the presence of SigB under osmotic stress conditions and SigH during morphological differentiation. Together with the phenotypic analysis of sigB and sigH operon mutants in S. coelicolor A3(2), the results suggest a dominant role for SigB in the osmotic stress response and a dual role for SigH in the osmotic stress response and morphological differentiation. These data suggest a complex regulation of the osmotic stress response in relation to morphological differentiation in S. coelicolor A3(2).


Asunto(s)
Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión Génica , Operón , Regiones Promotoras Genéticas , Factor sigma/genética , Streptomyces coelicolor/genética , Transcripción Genética , Proteínas Bacterianas/metabolismo , Secuencia de Bases , Factor sigma/metabolismo , Streptomyces coelicolor/crecimiento & desarrollo , Streptomyces coelicolor/metabolismo
19.
Front Cell Infect Microbiol ; 11: 692224, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34368013

RESUMEN

The chlamydial small non coding RNA, IhtA, regulates the expression of both HctA and DdbA, the uncharacterized product of the C. trachomatis L2 CTL0322 gene. HctA is a small, highly basic, DNA binding protein that is expressed late in development and mediates the condensation of the genome during RB to EB differentiation. DdbA is conserved throughout the chlamydial lineage, and is predicted to express a small, basic, cytoplasmic protein. As it is common for sRNAs to regulate multiple mRNAs within the same physiological pathway, we hypothesize that DdbA, like HctA, is involved in RB to EB differentiation. Here, we show that DdbA is a DNA binding protein, however unlike HctA, DdbA does not contribute to genome condensation but instead likely has nuclease activity. Using a DdbA temperature sensitive mutant, we show that DdbAts creates inclusions indistinguishable from WT L2 in size and that early RB replication is likewise similar at the nonpermissive temperature. However, the number of DdbAts infectious progeny is dramatically lower than WT L2 overall, although production of EBs is initiated at a similar time. The expression of a late gene reporter construct followed live at 40°C indicates that late gene expression is severely compromised in the DdbAts strain. Viability assays, both in host cells and in axenic media indicate that the DdbAts strain is defective in the maintenance of EB infectivity. Additionally, using Whole Genome Sequencing we demonstrate that chromosome condensation is temporally separated from DNA replication during the RB to EB transition. Although DdbA does not appear to be directly involved in this process, our data suggest it is a DNA binding protein that is important in the production and maintenance of infectivity of the EB, perhaps by contributing to the remodeling of the EB chromosome.


Asunto(s)
Chlamydia trachomatis , Regulación Bacteriana de la Expresión Génica , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Chlamydia trachomatis/genética , Chlamydia trachomatis/metabolismo , Proteínas de Unión al ADN/genética , Cuerpos de Inclusión/metabolismo
20.
Int J Mol Sci ; 22(15)2021 Aug 03.
Artículo en Inglés | MEDLINE | ID: mdl-34361108

RESUMEN

Alfalfa has emerged as one of the most important forage crops, owing to its wide adaptation and high biomass production worldwide. In the last decade, the emergence of bacterial stem blight (caused by Pseudomonas syringae pv. syringae ALF3) in alfalfa has caused around 50% yield losses in the United States. Studies are being conducted to decipher the roles of the key genes and pathways regulating the disease, but due to the sparse knowledge about the infection mechanisms of Pseudomonas, the development of resistant cultivars is hampered. The database alfaNET is an attempt to assist researchers by providing comprehensive Pseudomonas proteome annotations, as well as a host-pathogen interactome tool, which predicts the interactions between host and pathogen based on orthology. alfaNET is a user-friendly and efficient tool and includes other features such as subcellular localization annotations of pathogen proteins, gene ontology (GO) annotations, network visualization, and effector protein prediction. Users can also browse and search the database using particular keywords or proteins with a specific length. Additionally, the BLAST search tool enables the user to perform a homology sequence search against the alfalfa and Pseudomonas proteomes. With the successful implementation of these attributes, alfaNET will be a beneficial resource to the research community engaged in implementing molecular strategies to mitigate the disease. alfaNET is freely available for public use at http://bioinfo.usu.edu/alfanet/.


Asunto(s)
Proteínas Bacterianas/metabolismo , Bases de Datos de Proteínas , Interacciones Huésped-Patógeno , Medicago sativa/metabolismo , Enfermedades de las Plantas/inmunología , Mapas de Interacción de Proteínas , Pseudomonas syringae/patogenicidad , Medicago sativa/inmunología , Medicago sativa/microbiología , Enfermedades de las Plantas/microbiología
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...