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1.
Int J Mol Sci ; 22(19)2021 Sep 28.
Article in English | MEDLINE | ID: mdl-34638798

ABSTRACT

Detecting the folding/unfolding pathways of biological macromolecules is one of the urgent problems of molecular biophysics. The unfolding of bacterial luciferase from Vibrio harveyi is well-studied, unlike that of Photobacterium leiognathi, despite the fact that both of them are actively used as a reporter system. The aim of this study was to compare the conformational transitions of these luciferases from two different protein subfamilies during equilibrium unfolding with urea. Intrinsic steady-state and time-resolved fluorescence spectra and circular dichroism spectra were used to determine the stages of the protein unfolding. Molecular dynamics methods were applied to find the differences in the surroundings of tryptophans in both luciferases. We found that the unfolding pathway is the same for the studied luciferases. However, the results obtained indicate more stable tertiary and secondary structures of P. leiognathi luciferase as compared to enzyme from V. harveyi during the last stage of denaturation, including the unfolding of individual subunits. The distinctions in fluorescence of the two proteins are associated with differences in the structure of the C-terminal domain of α-subunits, which causes different quenching of tryptophan emissions. The time-resolved fluorescence technique proved to be a more effective method for studying protein unfolding than steady-state methods.


Subject(s)
Luciferases, Bacterial/chemistry , Molecular Dynamics Simulation , Photobacterium/chemistry , Vibrio/chemistry , Protein Domains , Spectrometry, Fluorescence
2.
Food Microbiol ; 99: 103679, 2021 Oct.
Article in English | MEDLINE | ID: mdl-34119089

ABSTRACT

Photobacterium spp. occur frequently in marine environments but have been recently also found as common spoilers on chilled meats. The environmental conditions in these ecological niches differ especially regarding salinity and ambient pressure. Linking the occurrence of photobacteria in different niches may elucidate its ecology and bring insights for the food industry. We investigated tolerance of Photobacterium (P.) phosphoreum and P. carnosum strains to high hydrostatic pressure and salinity and aligned our observations with presence of relevant genes. The strains were isolated from packaged meats and salmon (or the sea) to identify adaptations to marine and terrestrial habitats. Growth of all P. carnosum strains was reduced by 40 MPa hydrostatic pressure and >3% sodium chloride, suggesting loss of traits associated with marine habitats. In contrast, P. phosphoreum strains were only slightly affected, suggesting general adaptation to marine habitats. In accordance, these strains had gene clusters associated with marine niches, e.g. flagellar and lux-operons, being incomplete in P. carnosum. Occurrence of P. carnosum strains on packaged salmon and P. phosphoreum strains on meats therefore likely results from cross-contamination in meat and fish processing. Still, these strains showed intermediate traits regarding pressure- and halotolerance, suggesting developing adaptation to their respective environment.


Subject(s)
Meat/microbiology , Photobacterium/metabolism , Salmon/microbiology , Sodium Chloride/metabolism , Animals , Cattle , Chickens , Food Microbiology , Hydrostatic Pressure , Photobacterium/chemistry , Photobacterium/growth & development , Photobacterium/isolation & purification , Seawater/microbiology , Sodium Chloride/analysis
3.
Food Chem ; 309: 125691, 2020 Mar 30.
Article in English | MEDLINE | ID: mdl-31679853

ABSTRACT

This study presented a HPTLC platformed luminescent biosensor system for screening captan residue. First, the potential bio-effects of layers materials on the detectability of a luminescent bacteria Photobacterium phosphoreum (ATCC 11040) as the sensor cell were assessed. From comparison, it was noteworthy that the combination of sensor cells with normal silica gel layer exclusively gave outstanding detectability (<10 ng/zone). On this basis, HPTLC mediated separation and biosensing was further optimized. Then, the obtained graphic results were digitally quantified via software processing, offering satisfactory selectivity, linearity (R2 = 0.9901 within 10-80 ng/zone) and sensitivity (0.5 mg/kg against MRLs ≥ 6 mg/kg). Additionally, the performance of the established method was validated with different fruits (recover rates 75-96%, RSD < 11.8%). Meanwhile, it was demonstrated that detectability of this hybrid system would be tuneable by altering the combination of bacteria strains and layer materials, which was meaningful to strengthen the usability of microbial biosensors.


Subject(s)
Biosensing Techniques/methods , Captan/analysis , Fungicides, Industrial/analysis , Chromatography, Thin Layer , Fruit/chemistry , Fruit/metabolism , Malus/chemistry , Malus/metabolism , Photobacterium/chemistry , Photobacterium/isolation & purification , Silica Gel/chemistry
4.
Chem Commun (Camb) ; 55(52): 7458-7461, 2019 Jul 04.
Article in English | MEDLINE | ID: mdl-31184643

ABSTRACT

The development of a sensitive and reliable method for the detection of bioaccumulated heavy metal toxins is highly desirable for biotoxicity evaluation. However, the conventional biotoxicity evaluation method based on luminescent bacteria suffers from only being able to detect the overall toxicity without selectivity in light-off detection mode. Although various synthetic fluorescent probes have been developed for the selective detection of heavy metal ions, they usually suffer from aggregation-caused quenching after local accumulation in biological systems. To tackle these challenges, we herein develop a dual detection strategy for bioaccumulated Hg2+ based on turn-off of the bioluminescence of P. phosphoreum bacteria by disrupting the quorum sensing system and turn-on of the photoluminescence of an aggregation-induced emission (AIE) probe by forming aggregates with Hg2+ inside the bacteria. It is expected that the dual detection strategy would find broad applications in the evaluation of bioaccumulated toxins.


Subject(s)
Mercury/chemistry , Photobacterium/chemistry , Fluorescent Dyes/chemistry , Ions/chemistry , Light , Luminescent Measurements/methods , Mercury/pharmacology , Microscopy, Confocal , Photobacterium/isolation & purification , Quantum Theory , Quorum Sensing/drug effects
5.
Syst Appl Microbiol ; 41(5): 444-451, 2018 Sep.
Article in English | MEDLINE | ID: mdl-29804705

ABSTRACT

Three strains, H01100409BT, H01100413B, and H27100402HT, were isolated from several internal organs of diseased redbanded seabream (Pagrus auriga) reared in Andalusia (Southern Spain). All strains were studied by phenotypic, including chemotaxonomy, and genomic characteristics. Phylogenetic analysis based on concatenated sequences of six housekeeping genes (gyrB, ftsZ, topA, mreB, gapA, and 16S rRNA) supported the inclusion of the strains within the clade Phosphoreum of the genus Photobacterium, and two of the strains (H27100402HT and H01100409BT) formed a tight group separated from the closest species P. aquimaris. Genomic analyses, including average nucleotide identity (ANIb and ANIm) and DNA-DNA hybridization (DDH), clearly separated strains H27100402HT and H01100409BT from the other species within the clade Phosphoreum with values below the thresholds for species delineation. The chemotaxonomic features (including FAME analysis and MALDI-TOF-MS) of H27100402HT and H01100409BT strains confirmed their differentiation from the related taxa. The results demonstrated that strain H01100413B was classified as P. aquimaris and the strains H27100402HT and H01100409BT represented a new species each in the genus Photobacterium, for which we propose the names Photobacterium malacitanum sp. nov., type strain H27100402HT (=CECT 9190T=LMG 29992T), and Photobacterium andalusiense sp. nov., type strain H01100409BT (=CECT 9192T=LMG 29994T).


Subject(s)
Fish Diseases/microbiology , Photobacterium/classification , Photobacterium/physiology , Phylogeny , Animals , Base Composition , DNA, Bacterial/genetics , Fish Diseases/epidemiology , Fisheries , Genes, Bacterial/genetics , Genome, Bacterial/genetics , Phenotype , Photobacterium/chemistry , Photobacterium/genetics , RNA, Ribosomal, 16S/genetics , Sequence Analysis, DNA/veterinary , Spain/epidemiology , Species Specificity , Vitamin K 2/analogs & derivatives , Vitamin K 2/chemistry
6.
ACS Nano ; 11(12): 12840-12848, 2017 12 26.
Article in English | MEDLINE | ID: mdl-29195044

ABSTRACT

Biomimetic organization provides a promising strategy to develop functional materials and understand biological processes. However, how to mimic complex biological systems using simple biomolecular units remains a great challenge. Herein, we design and fabricate a biomimetic cyanobacteria model based on self-integration of small bioinspired molecules, including amphiphilic amino acid, 3,4-dihydroxyphenylalanine (DOPA), and metalloporphyrin and cobalt oxide nanoparticles (Co3O4 NPs), with the assistance of chemical conjugation and molecular self-assembly. The assembled amino acid fiber can be modified by DOPA to form covalently bound DOPA melanin containing hydroxyl and quinone species via Schiff base reaction. The adhering template can further tune the self-assembly of metalloporphyrin and Co3O4 NPs into J-aggregation and dispersive distribution, respectively, mainly via coordination binding. Metalloporphyrin molecules in the resulting hybrid fibers capture light; quinone species accept the excited electrons, and Co3O4 NPs catalyze water oxidation. Thus, the essential components of the photosystem-II protein complex in cyanobacteria are simplified and engineered into a simple framework, still retaining a similar photosynthetic mechanism. In addition, this architecture leads to efficient coupling of antenna, quinone-type reaction center, and photocatalyst, which increases the flux of light energy from antenna to reaction center for charge separation, resulting in enhanced oxygen evolution rate with excellent sustainability.


Subject(s)
Amino Acids/chemistry , Biomimetic Materials/chemistry , Cobalt/chemistry , Metalloporphyrins/chemistry , Oxides/chemistry , Oxygen/chemistry , Photobacterium/chemistry , Amino Acids/metabolism , Biomimetic Materials/metabolism , Cobalt/metabolism , Dihydroxyphenylalanine/chemistry , Dihydroxyphenylalanine/metabolism , Metalloporphyrins/metabolism , Molecular Structure , Oxides/metabolism , Oxygen/metabolism , Photobacterium/metabolism , Photosystem II Protein Complex/chemistry , Photosystem II Protein Complex/metabolism
7.
Luminescence ; 31(6): 1283-9, 2016 Sep.
Article in English | MEDLINE | ID: mdl-26864478

ABSTRACT

We demonstrated the possibility of long-term and efficient application of a biosensitive element (BE) in the form of Photobacterium phosphoreum photobacteria immobilized in poly(vinyl alcohol) (PVA) cryogel for detecting various ecotoxicants (Zn(2) (+) , Cu(2) (+) , Hg(2) (+) , Pb(2) (+) , 2,4-dichlorophenoxyacetic acid, 2,6-dimethylphenol, pentachlorophenol, coumaphos, malathion, chlorpyrifos and methyl parathion) in flow-through media. The range of detectable concentrations of ecotoxicants was determined at 1 × 10(-8) to 1 × 10(-4) M for heavy metal ions and at 1 × 10(-8) to 1 × 10(-5) M for phenol derivatives and organophosphorus pesticides. Immobilized cells of photobacteria quantitatively reacted with these ecotoxicants; cell sensitivity exhibited no flow rate dependence in the range from 45 to 180 mL/h. At a constant concentration of ecotoxicant in the flow, the bioluminescence quenching profile of immobilized cells demonstrated an integral response. The BE could remain in a flow-through medium for at least 10 days while retaining 95% of luminescent activity in the absence of ecotoxicants. The BE tested in this work was demonstrated to have a long shelf life (> 60 weeks) at -80°C without changes in the baseline level of bioluminescence. Copyright © 2016 John Wiley & Sons, Ltd.


Subject(s)
Biosensing Techniques , Cells, Immobilized/chemistry , Flow Injection Analysis , Luminescence , Photobacterium/chemistry , Water Pollutants, Chemical/analysis , 2,4-Dichlorophenoxyacetic Acid/analysis , Chlorpyrifos/analysis , Coumaphos/analysis , Malathion/analysis , Metals, Heavy/analysis , Methyl Parathion/analysis , Xylenes/analysis
8.
Biofizika ; 60(5): 898-905, 2015.
Article in Russian | MEDLINE | ID: mdl-26591600

ABSTRACT

The UV resistance of luminescent bacteria Escherichia coli AB1886 uvrA6 (pLeo1) containing the plasmid with luxCDABE genes of marine bacteria Photobacterium leiognathi is approximately two times higher than the UV resistance of non-luminous bacteria E. coli AB1886 uvrA6. Introduction of phr::kan(r) mutations (a defect in the functional activity of photolyase) into the genome of E. coli AB1886 uvrA6 (pLeo1) completely removes the high UV resistance of the cells. Therefore, photoreactivation that involves bacterial photolyase contributes mainly to the bioluminescence-induced DNA repair. It is shown that photoreactivating activity of bioluminescence of P. leiognathi is about 2.5 times lower compared with that one induced by a light source with λ > 385 nm. It is also shown that an increase in the bioluminescence intensity, induced by UV radiation in E. coli bacterial cells with a plasmid containing the luxCD ABE genes under RecA-LexA-regulated promoters, occurs only 25-30 min later after UV irradiation of cells and does not contribute to DNA repair. A quorum sensing regulatory system is not involved in the DNA repair by photolyase.


Subject(s)
DNA Damage/radiation effects , Escherichia coli/radiation effects , Photobacterium/chemistry , Ultraviolet Rays , DNA Damage/genetics , DNA Repair/genetics , Escherichia coli/genetics , Luminescence , Luminescent Proteins/chemistry , Mutation/radiation effects , Photobacterium/genetics , Promoter Regions, Genetic/radiation effects
9.
Biochemistry (Mosc) ; 80(6): 733-44, 2015 Jun.
Article in English | MEDLINE | ID: mdl-26531018

ABSTRACT

The scientific basis for producing luminescent biosensors containing free and immobilized luminescent bacteria is discussed. Modern technologies for engineering target objects, procedures used to immobilize bacteria in different carriers, as well as procedures for integral and specific biodetection of toxins are presented. Data regarding generation and application of biomonitoring for ecotoxicants derived from natural and genetically engineered photobacterial strains are analyzed. Special attention is given to immobilization of photobacteria in polyvinyl alcohol-containing cryogel. The main physicochemical, biochemical, and technological parameters for stabilizing luminescence in immobilized bacteria are described. Results of the application of immobilized photobacterial preparations both during discrete and continuous biomonitoring for different classes of ecotoxicants are presented.


Subject(s)
Biosensing Techniques/methods , Luminescent Measurements/methods , Luminescent Proteins/chemistry , Photobacterium/chemistry , Luminescent Proteins/metabolism , Photobacterium/metabolism
10.
Bull Exp Biol Med ; 159(5): 692-5, 2015 Sep.
Article in English | MEDLINE | ID: mdl-26468025

ABSTRACT

Bioluminescent method for measurements of the neutrophilic NAD(P)-dependent dehydrogenases (lactate dehydrogenase, NAD-dependent malate dehydrogenase, NADP-dependent decarboxylating malate dehydrogenase, NAD-dependent isocitrate dehydrogenase, and glucose- 6-phosphate dehydrogenase) is developed. The sensitivity of the method allows minimization of the volume of biological material for measurements to 104 neutrophils per analysis. The method is tried in patients with diffuse purulent peritonitis. Low levels of NADPH synthesis enzymes and high levels of enzymes determining the substrate flow by the Krebs cycle found in these patients can lead to attenuation of functional activity of cells.


Subject(s)
Biological Assay/standards , Glucosephosphate Dehydrogenase/metabolism , Isocitrate Dehydrogenase/metabolism , L-Lactate Dehydrogenase/metabolism , Malate Dehydrogenase (NADP+)/metabolism , Malate Dehydrogenase/metabolism , Peritonitis/diagnosis , FMN Reductase/chemistry , Humans , Hydrogen-Ion Concentration , Luciferases, Bacterial/chemistry , Luminescent Measurements , Neutrophils/chemistry , Neutrophils/enzymology , Neutrophils/pathology , Peritonitis/blood , Peritonitis/pathology , Photobacterium/chemistry , Photobacterium/enzymology , Reproducibility of Results , Sensitivity and Specificity
11.
J Bacteriol ; 197(23): 3676-85, 2015 Dec.
Article in English | MEDLINE | ID: mdl-26350139

ABSTRACT

UNLABELLED: Bacterial bioluminescence is taxonomically restricted to certain proteobacteria, many of which belong to the Vibrionaceae. In the most well-studied cases, pheromone signaling plays a key role in regulation of light production. However, previous reports have indicated that certain Photobacterium strains do not use this regulatory method for controlling luminescence. In this study, we combined genome sequencing with genetic approaches to characterize the regulation of luminescence in Photobacterium leiognathi strain KNH6, an extremely bright isolate. Using transposon mutagenesis and screening for decreased luminescence, we identified insertions in genes encoding components necessary for the luciferase reaction (lux, lum, and rib operons) as well as in nine other loci. These additional loci encode gene products predicted to be involved in the tricarboxylic acid (TCA) cycle, DNA and RNA metabolism, transcriptional regulation, and the synthesis of cytochrome c, peptidoglycan, and fatty acids. The mutagenesis screen did not identify any mutants with disruptions of predicted pheromone-related loci. Using targeted gene insertional disruptions, we demonstrate that under the growth conditions tested, luminescence levels do not appear to be controlled through canonical pheromone signaling systems in this strain. IMPORTANCE: Despite the long-standing interest in luminous bacteria, outside a few model organisms, little is known about the regulation and function of luminescence. Light-producing marine bacteria are widely distributed and have diverse lifestyles, suggesting that the control and significance of luminescence may be similarly diverse. In this study, we apply genetic tools to the study of regulation of light production in the extremely bright isolate Photobacterium leiognathi KNH6. Our results suggest an unusual lack of canonical pheromone-mediated control of luminescence and contribute to a better understanding of alternative strategies for regulation of a key bacterial behavior. These experiments lay the groundwork for further study of the regulation and role of bioluminescence in P. leiognathi.


Subject(s)
Bacterial Proteins/genetics , Photobacterium/chemistry , Photobacterium/genetics , Bacterial Proteins/metabolism , Gene Expression Regulation, Bacterial , Luciferases/genetics , Luciferases/metabolism , Luminescence , Molecular Sequence Data , Mutagenesis, Insertional , Operon , Photobacterium/enzymology , Photobacterium/metabolism
12.
Infect Immun ; 83(11): 4335-48, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26303391

ABSTRACT

Photobacterium damselae subsp. damselae, an important pathogen of marine animals, may also cause septicemia or hyperaggressive necrotizing fasciitis in humans. We previously showed that hemolysin genes are critical for virulence of this organism in mice and fish. In the present study, we characterized the hlyA gene product, a putative small ß-pore-forming toxin, and termed it phobalysin P (PhlyP), for "photobacterial lysin encoded on a plasmid." PhlyP formed stable oligomers and small membrane pores, causing efflux of K(+), with no significant leakage of lactate dehydrogenase but entry of vital dyes. The latter feature distinguished PhlyP from the related Vibrio cholerae cytolysin. Attack by PhlyP provoked a loss of cellular ATP, attenuated translation, and caused profound morphological changes in epithelial cells. In coculture experiments with epithelial cells, Photobacterium damselae subsp. damselae led to rapid hemolysin-dependent membrane permeabilization. Unexpectedly, hemolysins also promoted the association of P. damselae subsp. damselae with epithelial cells. The collective observations of this study suggest that membrane-damaging toxins commonly enhance bacterial adherence.


Subject(s)
Bacterial Toxins/metabolism , Hemolysin Proteins/metabolism , Photobacterium/metabolism , Amino Acid Sequence , Animals , Bacterial Adhesion , Bacterial Toxins/chemistry , Bacterial Toxins/genetics , Bacterial Toxins/toxicity , Epithelial Cells/microbiology , Erythrocytes/cytology , Erythrocytes/drug effects , Hemolysin Proteins/chemistry , Hemolysin Proteins/genetics , Hemolysin Proteins/toxicity , Hemolysis , Humans , Molecular Sequence Data , Photobacterium/chemistry , Photobacterium/genetics , Rabbits , Sequence Alignment
13.
Biochim Biophys Acta ; 1854(10 Pt A): 1466-75, 2015 Oct.
Article in English | MEDLINE | ID: mdl-26209460

ABSTRACT

The lux-operon of bioluminescent bacteria contains the genes coding for the enzymes required for light emission. Some species of Photobacteria feature an additional gene, luxF, which shows similarity to luxA and luxB, the genes encoding the heterodimeric luciferase. Isolated dimeric LuxF binds four molecules of an unusually derivatized flavin, i.e., 6-(3'-(R)-myristyl)-FMN (myrFMN). In the present study we have heterologously expressed LuxF in Escherichia coli BL21 in order to advance our understanding of the protein's binding properties and its role in photobacterial bioluminescence. Structure determination by X-ray crystallography confirmed that apo-LuxF possesses four preorganized binding sites, which are further optimized by adjusting the orientation of amino acid side chains. To investigate the binding properties of recombinant LuxF we have isolated myrFMN from Photobacterium leiognathi S1. We found that LuxF binds myrFMN tightly with a dissociation constant of 80±20 nM demonstrating that the purified apo-form of LuxF is fully competent in myrFMN binding. In contrast to LuxF, binding of myrFMN to luciferase is much weaker (Kd=4.0±0.4 µM) enabling LuxF to prevent inhibition of the enzyme by scavenging myrFMN. Moreover, we have used apo-LuxF to demonstrate that myrFMN occurs in all Photobacteria tested, irrespective of the presence of luxF indicating that LuxF is not required for myrFMN biosynthesis.


Subject(s)
Apoproteins/chemistry , Bacterial Proteins/chemistry , Flavin Mononucleotide/chemistry , Luciferases/chemistry , Myristic Acid/chemistry , Photobacterium/chemistry , Amino Acid Sequence , Apoproteins/genetics , Apoproteins/metabolism , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Crystallography, X-Ray , Escherichia coli/genetics , Escherichia coli/metabolism , Gene Expression , Luciferases/genetics , Luciferases/metabolism , Luminescence , Models, Molecular , Molecular Sequence Data , Photobacterium/enzymology , Protein Binding , Protein Conformation , Protein Multimerization , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Thermodynamics
14.
Biofizika ; 60(2): 316-21, 2015.
Article in Russian | MEDLINE | ID: mdl-26016027

ABSTRACT

This contribution presents the results of analysis of the dynamics of the bioluminescence of luminous bacteria Photobacterium phosphoreum IMV B-7071 under optimal conditions of their growth. A quasi-harmonic nature of the bacterial bioluminescence dynamics was detected. The observed periods of these changes have similar values compared with those in the earlier defined periods of changes in physicochemical properties of water. The relationship between biorhythms and a quasi-harmonic nature of changes in physicochemical properties of water is discussed.


Subject(s)
Luminescence , Photobacterium/chemistry , Water/chemistry , Luminescent Measurements , Periodicity , Photobacterium/growth & development
15.
J Nat Prod ; 78(3): 441-52, 2015 Mar 27.
Article in English | MEDLINE | ID: mdl-25699470

ABSTRACT

Sampling of California nearshore sediments resulted in the isolation of a Gram-negative bacterium, Photobacterium halotolerans, capable of producing unusual biosynthetic products. Liquid culture in artificial seawater-based media provided cyclic depsipeptides including four known compounds, kailuins B-E (2-5), and two new analogues, kailuins G and H (7 and 8). The structures of the new and known compounds were confirmed through extensive spectroscopic and Marfey's analyses. During the course of these studies, a correction was made to the previously reported double-bond geometry of kailuin D (4). Additionally, through the application of a combination of derivatization with Mosher's reagent and extensive (13)C NMR shift analysis, the previously unassigned chiral center at position C-3 of the ß-acyloxy group of all compounds was determined. To evaluate bioactivity and structure-activity relationships, the kailuin core (13) and kailuin lactam (14) were prepared by chiral synthesis using an Fmoc solid-phase peptide strategy followed by solution-phase cyclization. All isolated compounds and synthetic cores were assayed for solid tumor cell cytotoxicity and showed only minimal activity, contrary to other published reports. Additional phenotypic screenings were done on 4 and 5, with little evidence of activity.


Subject(s)
Biological Factors/chemistry , Biological Factors/isolation & purification , Depsipeptides/chemistry , Depsipeptides/isolation & purification , Gram-Negative Bacteria/chemistry , Photobacterium/chemistry , Humans , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Structure-Activity Relationship
16.
Org Lett ; 16(21): 5820-3, 2014 Nov 07.
Article in English | MEDLINE | ID: mdl-25343747

ABSTRACT

Prepiscibactin (1) is a possible intermediate in the biosynthesis of piscibactin, the siderophore responsible for the iron uptake of the bacterium Photobacterium damselae subsp. piscicida, the aethiological agent of fish pasteurellosis. Compound 1 was synthesized by a convergent approach starting from L-/D-cysteine and 2-hydroxybenzonitrile. The key steps were a highly diastereoselective SmI2-mediated Reformatsky reaction and Zn(2+)-induced asymmetric thiazolidine formation followed by lactamization. The absolute configuration 9R,10S,12R,13S was established for 1, and this confirmed the previous relative stereochemistry proposed on the basis of NOE and computational methods.


Subject(s)
Iodides/chemistry , Pfiesteria piscicida/chemistry , Photobacterium/chemistry , Samarium/chemistry , Thiazoles/chemistry , Thiazoles/chemical synthesis , Thiazolidines/chemistry , Zinc/chemistry , Animals , Cysteine/chemistry , Fish Diseases/microbiology , Fishes , Molecular Structure , Nitriles/chemistry , Pasteurella Infections/microbiology , Pasteurella Infections/pathology , Stereoisomerism
17.
J Vis Exp ; (87)2014 May 23.
Article in English | MEDLINE | ID: mdl-24894759

ABSTRACT

Fluorescence by Unbound Excitation from Luminescence (FUEL) is a radiative excitation-emission process that produces increased signal and contrast enhancement in vitro and in vivo. FUEL shares many of the same underlying principles as Bioluminescence Resonance Energy Transfer (BRET), yet greatly differs in the acceptable working distances between the luminescent source and the fluorescent entity. While BRET is effectively limited to a maximum of 2 times the Förster radius, commonly less than 14 nm, FUEL can occur at distances up to µm or even cm in the absence of an optical absorber. Here we expand upon the foundation and applicability of FUEL by reviewing the relevant principles behind the phenomenon and demonstrate its compatibility with a wide variety of fluorophores and fluorescent nanoparticles. Further, the utility of antibody-targeted FUEL is explored. The examples shown here provide evidence that FUEL can be utilized for applications where BRET is not possible, filling the spatial void that exists between BRET and traditional whole animal imaging.


Subject(s)
Fluorescence Resonance Energy Transfer/methods , Luminescent Measurements/methods , Escherichia coli/chemistry , Fluorescent Dyes/chemistry , Klebsiella pneumoniae/chemistry , Luciferases, Bacterial/chemistry , Nanoparticles/chemistry , Photobacterium/chemistry , Photobacterium/enzymology , Quantum Dots
18.
Aquat Toxicol ; 150: 165-74, 2014 May.
Article in English | MEDLINE | ID: mdl-24681700

ABSTRACT

The use of herbicide mixtures has become a cost-effective strategy against the evolution of herbicide resistance to protect global food production. Much research has focused on investigating either the herbicidal activities or the toxicity effects of herbicides; however, few of them have investigated both factors. This study investigates the balance between herbicidal activity for Selenastrum capricornutum and toxicity effect toward Photobacterium phosphoreum by determining the joint effects of triazine (simetryn, atrazine, prometon and prometryn) and phenylurea (fenuron, monuron, monolinuron and diuron) herbicides. The results showed that among the four triazines, only simetryn exhibited a unique effect (formation of a pi-sigma bond with the D1 microalga protein and an H-bond with the Luc photobacterial protein); and among 16 triazine-phenylurea binary mixtures, only the mixtures containing simetryn resulted in TU1 values (herbicidal activities of mixtures on S. capricornutum) >TU2 values (toxicity effects of mixtures on P. phosphoreum). However, the other 12 mixtures, which did not contain simetryn, showed the opposite result (TU1

Subject(s)
Chlorophyta/drug effects , Phenylurea Compounds/toxicity , Photobacterium/drug effects , Triazines/toxicity , Water Pollutants, Chemical/toxicity , Algal Proteins/chemistry , Algal Proteins/metabolism , Bacterial Proteins/chemistry , Bacterial Proteins/metabolism , Chlorophyta/chemistry , Herbicides/toxicity , Lethal Dose 50 , Models, Molecular , Photobacterium/chemistry
19.
PLoS One ; 9(1): e84992, 2014.
Article in English | MEDLINE | ID: mdl-24416329

ABSTRACT

Methicillin-resistant Staphylococcus aureus (MRSA) continues to be a serious human pathogen, and particularly the spread of community associated (CA)-MRSA strains such as USA300 is a concern, as these strains can cause severe infections in otherwise healthy adults. Recently, we reported that a cyclodepsipeptide termed Solonamide B isolated from the marine bacterium, Photobacterium halotolerans strongly reduces expression of RNAIII, the effector molecule of the agr quorum sensing system. Here we show that Solonamide B interferes with the binding of S. aureus autoinducing peptides (AIPs) to sensor histidine kinase, AgrC, of the agr two-component system. The hypervirulence of USA300 has been linked to increased expression of central virulence factors like α-hemolysin and the phenol soluble modulins (PSMs). Importantly, in strain USA300 Solonamide B dramatically reduced the activity of α-hemolysin and the transcription of psma encoding PSMs with an 80% reduction in toxicity of supernatants towards human neutrophils and rabbit erythrocytes. To our knowledge this is the first report of a compound produced naturally by a Gram-negative marine bacterium that interferes with agr and affects both RNAIII and AgrA controlled virulence gene expression in S. aureus.


Subject(s)
Gene Expression Regulation, Bacterial , Methicillin-Resistant Staphylococcus aureus/drug effects , Neutrophils/microbiology , Peptides, Cyclic/pharmacology , Quorum Sensing/drug effects , Animals , Bacterial Proteins/antagonists & inhibitors , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Bacterial Toxins/antagonists & inhibitors , Bacterial Toxins/genetics , Bacterial Toxins/metabolism , Erythrocytes/drug effects , Erythrocytes/microbiology , Hemolysin Proteins/antagonists & inhibitors , Hemolysin Proteins/genetics , Hemolysin Proteins/metabolism , Humans , Methicillin-Resistant Staphylococcus aureus/genetics , Methicillin-Resistant Staphylococcus aureus/pathogenicity , Neutrophils/drug effects , Peptides, Cyclic/isolation & purification , Photobacterium/chemistry , Primary Cell Culture , Protein Kinases/genetics , Protein Kinases/metabolism , Quorum Sensing/genetics , Rabbits , Trans-Activators/antagonists & inhibitors , Trans-Activators/genetics , Trans-Activators/metabolism , Virulence
20.
Prikl Biokhim Mikrobiol ; 50(5): 490-6, 2014.
Article in Russian | MEDLINE | ID: mdl-25707106

ABSTRACT

Immobilization of Photobacterium phosphoreum bacteria in polyvinyl alcohol cryogel was performed in order to develop biosensors used for ecotoxicant biomonitoring. The immobilization procedure, storage, and application of the immobilized cells for biomonitoring were optimized. It was shown that the immobilized cells demonstrate significantly higher stability and a longer duration of light emission than free bacteria. A discrete analysis of heavy metals and chlorophenols was conducted using the obtained biosensor samples.


Subject(s)
Biosensing Techniques/methods , Chlorophenols/analysis , Metals, Heavy/analysis , Photobacterium/chemistry , Water Pollutants, Chemical/analysis , Biosensing Techniques/instrumentation , Cells, Immobilized , Cryogels , Environmental Monitoring , Luminescent Measurements , Photobacterium/metabolism , Polyvinyl Alcohol/chemistry , Time Factors
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