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Journal of Inorganic Biochemistry Nov 2018A strategy for elucidating sequence determinants of function in the class of cytochrome P450 (CYP) enzymes that catalyze the first steps of terpene metabolism in wild...
A strategy for elucidating sequence determinants of function in the class of cytochrome P450 (CYP) enzymes that catalyze the first steps of terpene metabolism in wild microbiomes is described. Wild organisms that can use camphor, terpineol, pinene and limonene were isolated from soils rich in coniferous waste. Cell free extracts and growth beers were analyzed by gas chromatography/mass spectrometry to identify primary oxidative metabolites. For one organism, Pseudomonas nitroreducens TPJM, a cytochrome P450 (CYP108B1) isolated from cell free extracts was demonstrated to catalyze the oxidation of α-terpineol in assays combining the native ferredoxin and putidaredoxin reductase, and the resulting oxidation products identified by gas chromatography/mass spectrometry. Shotgun sequencing of PnTPJM identified four candidate P450 genes, including an apparently fragmentary gene with a high degree of homology with the known enzyme CYP108 (P450).
Topics: Bacterial Proteins; Cytochrome P-450 Enzyme System; Pseudomonas; Soil Microbiology; Terpenes
PubMed: 30170307
DOI: 10.1016/j.jinorgbio.2018.08.006 -
Canadian Journal of Microbiology Dec 2018We report an investigation of microbially induced carbonate precipitation by seven indigenous bacteria isolated from a landfill in China. Bacterial strains were cultured...
We report an investigation of microbially induced carbonate precipitation by seven indigenous bacteria isolated from a landfill in China. Bacterial strains were cultured in a medium supplemented with 25 mmol/L calcium chloride and 333 mmol/L urea. The experiments were carried out at 30 °C for 7 days with agitation by a shaking table at 130 r/min. Scanning electron microscopic and X-ray diffraction analyses showed variations in calcium carbonate polymorphs and mineral composition induced by all bacterial strains. The amount of carbonate precipitation was quantified by titration. The amount of carbonate precipitated in the medium varied among isolates, with the lowest being Bacillus aerius rawirorabr15 (LC092833) precipitating around 1.5 times more carbonate per unit volume than the abiotic (blank) solution. Pseudomonas nitroreducens szh_asesj15 (LC090854) was found to be the most efficient, precipitating 3.2 times more carbonate than the abiotic solution. Our results indicate that bacterial carbonate precipitation occurred through ureolysis and suggest that variations in carbonate crystal polymorphs and rates of precipitation were driven by strain-specific differences in urease expression and response to the alkaline environment. These results and the method applied provide benchmarking and screening data for assessing the bioremediation potential of indigenous bacteria for containment of contaminants in landfills.
Topics: Bacteria; Biomineralization; Calcium Carbonate; Crystallization; Hydrogen-Ion Concentration; Waste Disposal Facilities; X-Ray Diffraction
PubMed: 30148972
DOI: 10.1139/cjm-2018-0254 -
Biodegradation Jun 2018A bacterial strain NSA02, isolated from contaminated soil and identified as Pseudomonas nitroreducens based on partial 16S rDNA gene sequence analysis and BIOLOG...
A bacterial strain NSA02, isolated from contaminated soil and identified as Pseudomonas nitroreducens based on partial 16S rDNA gene sequence analysis and BIOLOG microbiology analysis, was used to study biodegradation of nicosulfuron in the culture medium. The optimal degradation conditions were determined to be 30 °C and pH 7.0. Batch tests were performed for seven different initial substrate concentrations to observe substrate degradation and associated cell growth. The biodegradation kinetics was found to follow a first-order model with regression values greater than 0.98. Specific degradation rate and specific growth rate of bacterial cells were observed to follow substrate inhibition kinetics, and the maximum values of both rates were observed at 100 mg L of nicosulfuron concentration. Kinetic parameters of three substrate inhibition models (Haldane, Aiba-Edwards and Teissier-Edwards) were fitted to the relationship between those rates and substrate concentrations. With the date obtained, Haldane and Teissier-Edwards models provide better representation when compared to Aiba-Edwards model. Inoculating nicosulfuron-treated soil samples with strain NSA02 resulted in a 5-6 times higher rate of nicosulfuron removal than that in non-inoculated soil. Five metabolites of nicosulfuron degradation were detected and identified by liquid chromatography mass spectrometry, and three possible biotransformation pathways were proposed. These results highlight the potential of the isolated bacterium to be used in the bioremediation of nicosulfuron-contaminated soils.
Topics: Biodegradation, Environmental; Biotransformation; DNA, Ribosomal; Hydrogen-Ion Concentration; Kinetics; Metabolome; Phylogeny; Pseudomonas; Pyridines; Soil Microbiology; Sulfonylurea Compounds; Temperature
PubMed: 29637426
DOI: 10.1007/s10532-018-9828-y -
Plant Cell Reports Jun 2018Pseudomonas nitroreducens: strain IHB B 13561 (PnIHB) enhances the growth of Arabidopsis thaliana and Lactuca sativa via the stimulation of cell development and nitrate...
Pseudomonas nitroreducens: strain IHB B 13561 (PnIHB) enhances the growth of Arabidopsis thaliana and Lactuca sativa via the stimulation of cell development and nitrate absorption. Plant growth-promoting rhizobacteria (PGPR) enhance plant development through various mechanisms; they improve the uptake of soil resources by plants to greatly promote plant growth. Here, we used Arabidopsis thaliana seedlings and Lactuca sativa to screen the growth enhancement activities of a purified PGPR, Pseudomonas nitroreducens strain IHB B 13561 (PnIHB). When cocultivated with PnIHB, both species of plants exhibited notably improved growth, particularly in regard to biomass. Quantitative reverse transcription polymerase chain reaction analysis indicated high expression levels of the nitrate transporter genes, especially NRT2.1, which plays a major role in the high-affinity nitrate transport system in roots. Moreover, enhanced activity of the cyclin-B1 promoter was observed when wild-type 'Columbia-0' Arabidopsis seedlings were exposed to PnIHB, whereas upregulation of cyclin-B also occurred in the inoculated lettuce seedlings. Overall, these results suggest that PnIHB improves A. thaliana and L. sativa growth via specific pathways involved in the promotion of cell development and enhancement of nitrate uptake.
Topics: Anion Transport Proteins; Arabidopsis; Biomass; Gene Expression Regulation, Plant; Lactuca; Nitrate Transporters; Nitrates; Plant Proteins; Plant Roots; Pseudomonas; Seedlings; Soil; Up-Regulation
PubMed: 29541882
DOI: 10.1007/s00299-018-2275-8 -
Genomics Data Dec 2017. DF05 is a Gram negative, motile, aerobic, rod-shaped and psychrotrophic bacterium that was isolated from contaminated San Jacinto River sediment near River Terrace...
. DF05 is a Gram negative, motile, aerobic, rod-shaped and psychrotrophic bacterium that was isolated from contaminated San Jacinto River sediment near River Terrace Park in Channelview, Texas. The draft genome of strain DF05 consists of a total of 192 contigs assembled at the scaffold level totaling 6,487,527 bp and encoding for 5862 functional proteins, 1116 of which are annotated as hypothetical proteins. The bacterial chromosome has a GC content of 65.15% and contains 22 rRNA and 70 tRNA loci. In addition, approximately 142 proteins localized on the bacterial chromosome are associated with metabolism of aromatic compounds. A single plasmid approximately 95 kb in size was also detected carrying copies of RNA genes and multiple phage assembly proteins.
PubMed: 28856100
DOI: 10.1016/j.gdata.2017.07.011 -
Applied Microbiology and Biotechnology Jun 2017The surfactant sodium lauryl ether sulfate (SLES) is widely used in the composition of detergents and frequently ends up in wastewater treatment plants (WWTPs). While...
The surfactant sodium lauryl ether sulfate (SLES) is widely used in the composition of detergents and frequently ends up in wastewater treatment plants (WWTPs). While aerobic SLES degradation is well studied, little is known about the fate of this compound in anoxic environments, such as denitrification tanks of WWTPs, nor about the bacteria involved in the anoxic biodegradation. Here, we used SLES as sole carbon and energy source, at concentrations ranging from 50 to 1000 mg L, to enrich and isolate nitrate-reducing bacteria from activated sludge of a WWTP with the anaerobic-anoxic-oxic (A/O) concept. In the 50 mg L enrichment, Comamonas (50%), Pseudomonas (24%), and Alicycliphilus (12%) were present at higher relative abundance, while Pseudomonas (53%) became dominant in the 1000 mg L enrichment. Aeromonas hydrophila strain S7, Pseudomonas stutzeri strain S8, and Pseudomonas nitroreducens strain S11 were isolated from the enriched cultures. Under denitrifying conditions, strains S8 and S11 degraded 500 mg L SLES in less than 1 day, while strain S7 required more than 6 days. Strains S8 and S11 also showed a remarkable resistance to SLES, being able to grow and reduce nitrate with SLES concentrations up to 40 g L. Strain S11 turned out to be the best anoxic SLES degrader, degrading up to 41% of 500 mg L. The comparison between SLES anoxic and oxic degradation by strain S11 revealed differences in SLES cleavage, degradation, and sulfate accumulation; both ester and ether cleavage were probably employed in SLES anoxic degradation by strain S11.
Topics: Aeromonas; Biodegradation, Environmental; Carbon; Comamonadaceae; Comamonas; Denitrification; Gram-Negative Bacteria; Oxidation-Reduction; Pseudomonas; Sewage; Sodium Dodecyl Sulfate; Surface-Active Agents
PubMed: 28299401
DOI: 10.1007/s00253-017-8212-x -
Applied and Environmental Microbiology Nov 2016Pseudomonas nitroreducens TX1 is of special interest because of its ability to utilize 0.05% to 20% octylphenol polyethoxylates (OPEO) as a sole source of carbon. In...
Pseudomonas nitroreducens TX1 is of special interest because of its ability to utilize 0.05% to 20% octylphenol polyethoxylates (OPEO) as a sole source of carbon. In this study, a library containing 30,000 Tn5-insertion mutants of the wild-type strain TX1 was constructed and screened for OPEO utilization, and 93 mutants were found to be unable to grow on OPEO In total, 42 separate disrupted genes were identified, and the proteins encoded by the genes were then classified into various categories, namely, information storage and processing (14.3%), cellular processes and signaling (28.6%), metabolism (35.7%), and unknown proteins (21.4%). The individual deletion of genes encoding isocitrate lyase (aceA), malate synthase (aceB), and glycolate dehydrogenase (glcE) was carried out, and the requirement for aceA and aceB but not glcE confirmed the role of the glyoxylate cycle in OPEO degradation. Furthermore, acetaldehyde dehydrogenase and acetyl-coenzyme A (acetyl-CoA) synthetase activity levels were 13.2- and 2.1-fold higher in TX1 cells grown on OPEO than in TX1 cells grown on succinate, respectively. Growth of the various mutants on different carbon sources was tested, and based on these findings, a mechanism involving exoscission to liberate acetaldehyde from the end of the OPEO chain during degradation is proposed for the breakdown of OPEO IMPORTANCE: Octylphenol polyethoxylates belong to the alkylphenol polyethoxylate (APEO) nonionic surfactant family. Evidence based on the analysis of intermediate metabolites suggested that the primary biodegradation of APEO can be achieved by two possible pathways for the stepwise removal of the C ethoxylate units from the end of the chain. However, direct evidence for these hypotheses is still lacking. In this study, we described the use of transposon mutagenesis to identify genes critical to the catabolism of OPEO by P. nitroreducens TX1. The exoscission of the ethoxylate chain leading to the liberation of acetaldehyde is proposed. Isocitrate lyase and malate synthase in glyoxylate cycle are required in the catabolism of ethoxylated surfactants. Our findings also provide many gene candidates that may help elucidate the mechanisms in stress responses to ethoxylated surfactants by bacteria.
Topics: Acetaldehyde; Alcohol Oxidoreductases; Aldehyde Oxidoreductases; Bacterial Proteins; DNA Transposable Elements; Gene Deletion; Isocitrate Lyase; Malate Synthase; Mutagenesis, Insertional; Phenols; Pseudomonas; Succinic Acid; Surface-Active Agents
PubMed: 27590807
DOI: 10.1128/AEM.01907-16 -
Journal of Hazardous Materials Jan 2017Effect of biosurfactant on biodegradation of pyrene was studied using a microbial consortium predominantly composed of Pseudomonas viridiflava (49.5%) and Pseudomonas...
Effect of biosurfactant on biodegradation of pyrene was studied using a microbial consortium predominantly composed of Pseudomonas viridiflava (49.5%) and Pseudomonas nitroreducens (32.5%) in a batch experiment containing lipopeptidic biosurfactant, produced by Paenibacillus dendritiformis CN5 strain, and mineral salt medium. The results showed that the lipopeptide at 600 and 300mgL enhanced pyrene degradation to 83.5% and 67% respectively in 24days compared to 16% degradation in its absence. However degradation of pyrene was reduced to 57% as the lipopeptide supplementation was raised to 900mgL. This demonstrates that the biodegradation of pyrene was found to increase with an increase in the lipopeptide concentration up to a threshold level. The experimental data were fitted to the logistic kinetic model which provided best fit with a coefficient of determination (R) values≥0.97. Maximum specific growth rate, μ of 0.97 and 0.69d were achieved in the 600 and 300mgL lipopeptide amendments in comparison to 0.54d in the unamended one. The carrying capacity, X increased 4.4-fold in 600mgL lipopeptide supplemented samples in comparison to its absence. Generally the lipopeptide showed potential application in improving bioremediation of polycyclic aromatic hydrocarbons contaminated environmental media.
Topics: Biodegradation, Environmental; Environmental Pollutants; Lipopeptides; Microbial Consortia; Paenibacillus; Pseudomonas; Pyrenes; Surface-Active Agents
PubMed: 27627697
DOI: 10.1016/j.jhazmat.2016.08.035 -
Applied Biochemistry and Biotechnology Apr 2015One clone exhibiting lipolytic activity was selected among 30 positives from a metagenomic library of a microbe consortium specialized in petroleum hydrocarbon...
One clone exhibiting lipolytic activity was selected among 30 positives from a metagenomic library of a microbe consortium specialized in petroleum hydrocarbon degradation. From this clone, a sublibrary was constructed and a metagenome contig was assembled and analyzed using the ORF Finder; thus, it was possible to identify a potential ORF that encodes a lipolytic enzyme, denoted ORF2. This ORF is composed of 1035-bp 345 amino acids and displayed 98 % identity with an alpha/beta hydrolase from Pseudomonas nitroreducens (accession number WP024765380.1). When analyzed against a metagenome database, ORF2 also showed 76 % of sequence identity with a hypothetical protein from a marine metagenome (accession number ECT55726.1). The ProtParam analyses indicated that the recombinant protein ORF2 has a molecular mass approximately 39 kDa, as expected from its amino acid sequence, and based on phylogenetic analysis and molecular modeling, it was possible to suggest that ORF2 is a new member from family V. This enzyme exhibits the catalytic triad and conserved motifs typical from this family, wherein the serine residue is located in the central position of the conserved motif GASMGG. The orf2 gene was cloned in the expression vector pET28a, and the recombinant protein was superexpressed in Escherichia coli BL21(DE3) cells. The lipolytic activity of protein bands presented in a SDS-PAGE gel was confirmed by zymogram analyses, indicating ORF2 activity. These discoveries raise the possibility of employing this protein in biotechnological applications, such as bioremediation.
Topics: Amino Acid Sequence; Bacterial Proteins; Cloning, Molecular; Gene Expression Regulation, Bacterial; Metagenome; Metagenomics; Models, Molecular; Petroleum; Phylogeny; Pseudomonas; Sequence Alignment; Substrate Specificity
PubMed: 25764223
DOI: 10.1007/s12010-015-1556-8 -
Pyrimidine nucleotide synthesis in Pseudomonas nitroreducens and the regulatory role of pyrimidines.Microbiological Research Dec 2014Control of pyrimidine biosynthesis in the commercially important, hydrocarbon-utilizing bacterium Pseudomonas nitroreducens ATCC 33634 was investigated. When...
Control of pyrimidine biosynthesis in the commercially important, hydrocarbon-utilizing bacterium Pseudomonas nitroreducens ATCC 33634 was investigated. When glucose-grown wild-type cells were supplemented with uracil or orotic acid, the pyrimidine biosynthetic activities were depressed. Pyrimidine limitation of glucose-grown cells of an orotate phosphoribosyltransferase mutant caused aspartate transcarbamoylase and dihydroorotase activities to increase by about 4-fold while the other enzyme activities about doubled. In succinate-grown phosphoribosyltransferase mutant cells subjected to pyrimidine limitation, transcarbamoylase and dehydrogenase activities rose by about 5-fold while dihydroorotase activity more than tripled. In an OMP decarboxylase mutant, pyrimidine limitation of glucose-grown cells increased transcarbamoylase, dihydroorotase, dehydrogenase and phosphoribosyltransferase activities by 4-, 10-, 6- and 3.8-fold, respectively. Pyrimidine limitation of the succinate-grown decarboxylase mutant cells increased aspartate transcarbamoylase or dihydroorotase by more than 4-fold and the other activities by about 2-fold. Pyrimidine biosynthetic enzyme synthesis appeared to be regulated by pyrimidines with the regulation being influenced by the carbon source present. Aspartate transcarbamoylase activity in Ps. nitroreducens was regulated at the level of enzyme activity since the enzyme was strongly inhibited by UDP, pyrophosphate, ATP and ADP. Overall, the regulation of pyrimidine biosynthesis in Ps. nitroreducens can be used to differentiate it from other taxonomically related species of Pseudomonas.
Topics: Aspartate Carbamoyltransferase; Pseudomonas; Pyrimidine Nucleotides; Pyrimidines
PubMed: 24867376
DOI: 10.1016/j.micres.2014.04.003