Purification and properties of pantothenase from Pseudomonas fluorescens.
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ABSTRACT:
SUBMITTER: Airas RK
PROVIDER: S-EPMC1163867 | biostudies-other | 1976 Aug
REPOSITORIES: biostudies-other
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Project description:1. l-Mandelate dehydrogenase and mandelate racemase were partially purified from extracts of Pseudomonas fluorescens A-312 grown in media containing d-mandelate. 2. The activity of mandelate racemase, but not that of l-mandelate dehydrogenase, is greatly stimulated by Mg(2+), Mn(2+), Co(2+) and, though less effectively, by Ni(2+). Other metal ions are inactive or inhibitory. 3. Racemase activity is inhibited by phosphate, fluoride, pyrophosphate and EDTA. The inhibitions by pyrophosphate and EDTA are competitive with respect to the metal ion activator; those by phosphate and fluoride are competitive with respect to the substrate. 4. The addition of Mg(2+) diminishes the Michaelis constant of racemase. 5. The pH optimum of the racemase is at 7.8. The pH-activity curve of the dehydrogenase complex of enzymes has two peaks, at 7.0 and 8.2. 6. The enzymic racemization of d-mandelate is initially faster than that of l-mandelate. 7. The rates of oxidation of related substrates, catalysed by l-mandelate dehydrogenase, are in the decreasing order: l-p-hydroxymandelate; l-3,4-dihydroxymandelate; l-4-hydroxy-3-methoxymandelate. The racemase is active towards d-p-hydroxymandelate but inactive towards d-3,4-dihydroxymandelate and d-4-hydroxy-3-methoxymandelate. Since 4-hydroxy-3-methoxymandelate, and presumably also 3,4-dihydroxymandelate, arising from the metabolism of catechol-amines, have the d-configuration, the enzymes studied cannot be utilized for estimation of the last two acids in urine.
| S-EPMC1270079 | biostudies-other
Project description:Zoospores play an important role in the infection of plant and animal hosts by oomycetes and other zoosporic fungi. In this study, six fluorescent Pseudomonas isolates with zoosporicidal activities were obtained from the wheat rhizosphere. Zoospores of multiple oomycetes, including Pythium species, Albugo candida, and Phytophthora infestans, were rendered immotile within 30 s of exposure to cell suspensions or cell culture supernatants of the six isolates, and subsequent lysis occurred within 60 s. The representative strain SS101, identified as Pseudomonas fluorescens biovar II, reduced the surface tension of water from 73 to 30 mN m-1. The application of cell suspensions of strain SS101 to soil or hyacinth bulbs provided significant protection against root rot caused by Pythium intermedium. Five Tn5 mutants of strain SS101lacked the abilities to reduce the surface tension of water and to cause lysis of zoospores. Genetic characterization of two surfactant-deficient mutants showed that the transposons had integrated into condensation domains of peptide synthetases. A partially purified extract from strain SS101 reduced the surface tension of water to 30 mN m-1 and reached the critical micelle concentration at 25 micrograms ml-1. Reverse-phase high-performance liquid chromatography yielded eight different fractions, five of which had surface activity and caused lysis of zoospores. Mass spectrometry and nuclear magnetic resonance analyses allowed the identification of the main constituent as a cyclic lipopeptide (1,139 Da) containing nine amino acids and a 10-carbon hydroxy fatty acid. The other four zoosporicidal fractions were closely related to the main constituent, with molecular massesranging from 1,111 to 1,169 Da.
| S-EPMC309978 | biostudies-literature
Project description:BackgroundPseudomonas fluorescens SBW25 has been extensively studied because of its plant growth promoting properties and potential as a biocontrol agent. The genome of SBW25 has been sequenced, and among sequenced strains of pseudomonads, SBW25 appears to be most closely related to P. fluorescens WH6. In the authors' laboratories, WH6 was previously shown to produce and secrete 4-formylaminooxyvinylglycine (FVG), a non-proteinogenic amino acid with selective herbicidal and antimicrobial activity. Although SBW25 does not have the genetic capacity to produce FVG, we were interested in determining whether this pseudomonad might produce some other type of non-proteinogenic amino acid.ResultsP. fluorescens SBW25 was found to produce and secrete a ninhydrin-reactive compound with selective antimicrobial properties. This compound was purified from SBW25 culture filtrate and identified as the non-proteinogenic amino acid L-furanomycin [2S,2'R,5'S)-2-amino-2-(5'methyl-2',5'-dihydrofuran-2'-yl)acetic acid].ConclusionsThe identification of furanomycin as a secondary metabolite of SBW25 is the first report of the production of furanomycin by a pseudomonad. This compound was known previously only as a natural product produced by a strain of Streptomyces. This report adds furanomycin to the small list of non-proteinogenic amino acids that have been identified as secondary products of pseudomonads. This study also extends the list of bacteria that are inhibited by furanomycin to include several plant pathogenic bacteria.
| S-EPMC3662646 | biostudies-literature
Project description:Massetolide A is a cyclic lipopeptide (CLP) antibiotic produced by various Pseudomonas strains from diverse environments. Cloning, sequencing, site-directed mutagenesis, and complementation showed that massetolide A biosynthesis in P. fluorescens SS101 is governed by three nonribosomal peptide synthetase (NRPS) genes, designated massA, massB, and massC, spanning approximately 30 kb. Prediction of the nature and configuration of the amino acids by in silico analysis of adenylation and condensation domains of the NRPSs was consistent with the chemically determined structure of the peptide moiety of massetolide A. Structural analysis of massetolide A derivatives produced by SS101 indicated that most of the variations in the peptide moiety occur at amino acid positions 4 and 9. Regions flanking the mass genes contained several genes found in other Pseudomonas CLP biosynthesis clusters, which encode LuxR-type transcriptional regulators, ABC transporters, and an RND-like outer membrane protein. In contrast to most Pseudomonas CLP gene clusters known to date, the mass genes are not physically linked but are organized in two separate clusters, with massA disconnected from massB and massC. Quantitative real-time PCR analysis indicated that transcription of massC is strongly reduced when massB is mutated, suggesting that these two genes function in an operon, whereas transcription of massA is independent of massBC and vice versa. Massetolide A is produced in the early exponential growth phase, and biosynthesis appears not to be regulated by N-acylhomoserine lactone-based quorum sensing. Massetolide A production is essential in swarming motility of P. fluorescens SS101 and plays an important role in biofilm formation.
| S-EPMC2293227 | biostudies-literature
Project description:In recent years, evolutionary biologists have developed an increasing interest in the use of barcoding strategies to study eco-evolutionary dynamics of lineages within evolving populations and communities. Although barcoded populations can deliver unprecedented insight into evolutionary change, barcoding microbes presents specific technical challenges. Here, strategies are described for barcoding populations of the model bacterium Pseudomonas fluorescens SBW25, including the design and cloning of barcoded regions, preparation of libraries for amplicon sequencing, and quantification of resulting barcoded lineages. In so doing, we hope to aid the design and implementation of barcoding methodologies in a broad range of model and non-model organisms.
| S-EPMC10275814 | biostudies-literature
Project description:The Mn(2+)-oxidizing bacterium Pseudomonas fluorescens GB-1 deposits Mn oxide around the cell. During growth of a culture, the Mn(2+)-oxidizing activity of the cells first appeared in the early stationary growth phase. It depended on the O2 concentration in the culture during the late logarithmic growth phase. Maximal activity was observed at an oxygen concentration of 26% saturation. The activity could be recovered in cell extracts and was proportional to the protein concentration in the cell extracts. The specific activity was increased 125-fold by ammonium sulfate precipitation followed by reversed-phase and gel filtration column chromatographies. The activity of the partly purified Mn(2+)-oxidizing preparation had a pH optimum of circa 7 and a temperature optimum of 35 degrees C and was lost by heating. The Mn(2+)-oxidizing activity was sensitive to NaN3 and HgCl2. It was inhibited by KCN, EDTA, Tris, and o-phenanthroline. Although most data indicated the involvement of protein in Mn2+ oxidation, the activity was slightly stimulated by sodium dodecyl sulfate at a low concentration and by treatment with pronase and V8 protease. By polyacrylamide gel electrophoresis, two Mn(2+)-oxidizing factors with estimated molecular weights of 180,000 and 250,000 were detected.
| S-EPMC168802 | biostudies-other
Project description:We find out the antimicrobial potential of partially purified 2,4-diacetylphloroglucinol (DAPG) against Ralstonia solanacearum and fungal plant pathogens isolated from tomato rhizobacterium Pseudomonas fluorescens VSMKU3054. The present study is mainly focused on the control of wilt disease of tomato by our isolate VSMKU3054 and DAPG. The cell free culture filtrate of P. fluorescens VSMKU3054 was significantly arrested the growth of R. solanacearum and fungal pathogens such as Rhizoctonia solani, Sclerotium rolfsii, Macrophomina phaseolina and Fusarium oxysporum compared to control. The existence of DAPG from the crude metabolites of P. fluorescens VSMKU3054 was confirmed on TLC with Rf value 0.34, which is coincide with that of authentic phloroglucinol. The partially purified DAPG exhibited much higher activity against R. solanacearum at 30 µg/ml than the fungal plant pathogens compared to control. The antimicrobial partially purified compound was identified as DAPG by UV, FT-IR and GC-MS analysis. The percentage of live cells of R. solanacearum when supplemented with DAPG at 30 µg/ml, significantly controlled the living nature of R. solanacearum up to 68% compared to tetracycline and universal control observed under high content screening analysis. The selected isolate P. fluorescens VSMKU3054 and DAPG significantly controlled wilt disease of tomato up to 59.5% and 42.12% on 3rd and 7th days compared to positive and negative control by detached leaf assay. Further, in silico analysis revealed that high interaction of DAPG encoding protease with lectin which is associated with R. solanacearum. Based on our findings, we confirmed that P. fluorescens VSMKU3054 and DAPG could be used a potential bio inoculants for the management of bacterial wilt disease of tomato.
| S-EPMC8071909 | biostudies-literature
Project description:The Pseudomonas aeruginosa genome encodes a variety of different proteolytic enzymes several of which play an important role as virulence factors. Interestingly, only two of these proteases are predicted to belong to the subtilase family and we have recently studied the physiological role of the subtilase SprP. Here, we describe the functional overexpression of SprP in Escherichia coli using a novel expression and secretion system. We show that SprP is autocatalytically activated by proteolysis and exhibits optimal activity at 50°C in a pH range of 7-8. We also demonstrate a significant increase in sprP promoter activity upon growth of P. aeruginosa at 43°C indicating a role for SprP in heat shock response.
| S-EPMC4618607 | biostudies-literature
Project description:The goal of the study was to examine the influence of flowback water exposure on bacterial survival and biocide tolerance. The transcriptome of P. fluorescens was analyzed using RNA-seq to determine the gene rxpression profile changes occuring upon flowback water exposure. The results indicate that that P. fluorescens induces a well-coordinated genetic response that aids in its survival in flowback water as well as imparts enhanced tolerance against typically used slow acting biocides such as gluteraldehyde but increased susceptibity towards sodium hypochlorite. RNA-seq data demonstrated significant induction of genes involved in osmotic stress, energy production and conversion, membrane integrity, protein transport among others.
2014-12-23 | GSE55388 | GEO
Project description:We report a genome update for Pseudomonas fluorescens isolate SBW25. The updated genome assembly, which was derived from the original isolate, is based on PacBio long-read sequence data. It shows three minor differences, compared with the previously published genome sequence. Original annotations were merged with recent automated annotations to preserve information.
| S-EPMC9933645 | biostudies-literature