• Microbiology and Biotechnology Letters
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• v.32 no.3
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• pp.206-211
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• 2004

Levansucrase gene(lscA) from Pseudomonas aurantiaca was subcloned downstream of GAL1 promoter in pYES 2.0 and pYInu-AT [GAL10 promoter ＋ exoinulinase signal sequence of Kluyveromyces marxianus], resulting pYES-lscA and p YInu-lscA, respectively. The two expression plasmids were introduced into an invertase-deficient strain, Sacchromayces cerevisiae SEY2102, and transformants with high activity of levansucrase were selected. When each yeast transform ants was cultivated in medium containing galactose, the extracellular and intracellular activities of levansucrase reached about 8.62 U/ml with the strain harboring pYES-lscA and 5.43 U/ml with the strain harboring pYInu-lscA. The levansucrase activity of 80% was detected in the periplasmic space and cytoplasm. The levansucrase activity in the medium of SEY2102/pYInu-lscA was 0.87 U/ml whereas that of SEY2102/pYES-lscA was 0.47 U/ml, which implying the exoinulinase signal sequence didn't enhance the secretion efficiency of levansucrase. Furthermore, the recombinant levansucrase expressed in yeast seems to be produced as a hyper-glycosylated form.

• Journal of Microbiology and Biotechnology
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• v.16 no.1
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• pp.102-108
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• 2006

The IscA gene, encoding a levansucrase of 424 amino acids (aa) residues, was cloned from the genomic DNA of Pseudomonas aurantiaca S-4380, and overexpressed in Escherichia coli. The recombinant levansucrase overexpressed in E. coli was then used to produce levan from sucrose. Levan crystals with 98% purity could be obtained from the reaction mixture with $62\%$ yield using an alcohol precipitation method. The molecular weight of the levan was $7\times10^5$ daltons. Methylation studies showed that the levan was branched: main linkage C-2,6; branched linkage C-2,1; and degree of branching $6\%$. Three bacterial levans from different strains were incubated with levan fructotransferase (LFTase) from Arthrobacter ureafaciens K2032, which produced $di-\beta-D-fructofuranose$ dianhydride IV (DFA IV); final conversion yields from the levans to DFA IV were $39\%$ in Zymomonas mobilis, $53\%$ in Serratia levanicum, and $59\%$ in P. aurantiaca S-4380 levansucrase. The levan from P. aurantiaca S-4380 levansucrase gave the highest conversion yield of levan to DFAIV so far reported.

• Journal of Microbiology and Biotechnology
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• v.27 no.3
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• pp.480-491
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• 2017

Fluorescent pseudomonads have been isolated from halophytes, mesophytes, and xerophytes of Pakistan. Among these, eight isolates, GS-1, GS-3, GS-4, GS-6, GS-7, FS-2 (cactus), ARS-38 (cotton), and RP-4 (para grass), showed antifungal activity and were selected for detailed study. Based on biochemical tests and 16S rRNA gene sequences, these were identified as strains of P. chlororaphis subsp. chlororaphis and aurantiaca. Secondary metabolites of these strains were analyzed by LC-MS. Phenazine-1-carboxylic acid (PCA), 2-hydroxy-phenazine, Cyclic Lipopeptide (white line-inducing principle (WLIP)), and lahorenoic acid A were detected in variable amounts in these strains. P. aurantiaca PB-St2 was used as a reference as it is known for the production of these compounds. The phzO and PCA genes were amplified to assure that production of these compounds is not an artifact. Indole acetic acid production was confirmed and quantified by HPLC. HCN and siderophore production by all strains was observed by plate assays. These strains did not solubilize phosphate, but five strains were positive for zinc solubilization. Wheat seedlings were inoculated with these strains to observe their effect on plant growth. P. aurantiaca strains PB-St2 and GS-6 and P. chlororaphis RP-4 significantly increased both root and shoot dry weights, as compared with uninoculated plants. However, P. aurantiaca strains FS-2 and ARS-38 significantly increased root and shoot dry weights, respectively. All strains except PB-St2 and ARS-38 significantly increased the root length. This is the first report of the isolation of P. aurantiaca from cotton and cactus, P. chlororaphis from para grass, WLIP and lahorenoic acid A production by P. chlororaphis, and zinc solubilization by P. chlororaphis and P. aurantiaca.

• Journal of Microbiology and Biotechnology
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• v.22 no.3
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• pp.326-330
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• 2012

Antifungal metabolites were isolated from a culture of Pseudomonas aurantiaca IB5-10. Chemical structures of the metabolites were elucidated as phenazine-1-carboxylic acid (PCA; 1), 2-hydroxyphenazine (2-OH-PHZ; 2), and cyclo-(L-Pro-L-Val; 3), respectively, based on spectroscopic methods. Among them, 3 was isolated for the first time from this strain. The antifungal activities of 1-3 were evaluated against a variety of plant pathogens. To the best of our knowledge, the antifungal activities of 3 against plant fungal pathogens have been evaluated for the first time in this work. PCA (1) showed the most potent antifungal activities against Phytophthora capsici, Rhizoctonia solani AG-1(IA), and Pythium ultimum with MICs (${\mu}g/ml$) of less than 1.0, 1.3, and 2.0, respectively. On the other hand, 2-OH-PHZ (2) showed potent antifungal activity against R. solani AG-1(IA) with the MIC (${\mu}g/ml$) of 2.0, whereas it showed moderate antifungal activity against P. ultimum with the MIC (${\mu}g/ml$) of 50.0. In addition, 3 showed antifungal activity against only R. solani AG-1(IA).

• KSBB Journal
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• v.18 no.4
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• pp.312-317
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• 2003

The addition of glycine up to 0.5％ (w/v) to Luria broth (LB) media on the secretion of levansucrase in a recombinant strain Escherichia coli JM109/pUPLK1 was observed to enhance the release of periplasmic proteins from the cell to the broth, without significantly affecting the cell growth rate and protein productivity. However, the glycine concentration at 1 ％ (w/v), the cell density attainable at the stationary phase fell to about 50％ and the extracellular activity of levansucrase corresponded to about 80％ of the total (extracellular plus intracellular) activity and increased by 2.6-fold, comparing to the cells grown in the absence of glycine. The increased pH at stationary phase accelerated the degradation of levansucrase. Maximal extracellular activity was attained when 1 ％ glycine was supplemented at the onset of strain growth.

• Journal of Microbiology and Biotechnology
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• v.12 no.4
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• pp.603-609
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• 2002

DFA IV is di-D-fructose-2,6':6,2'-dianhydride, consisting of two fructose residues. It can be enzymatically synthesized from levan by levan fructotransferase, and can be used for mineral absorption. Understanding of the structure and composition of levan is important to obtain high-level production of DFA IV. A bacterial strain, Pseudomonas aurantiaca 5-4380, was identified to produce low-branched levan, and the levansucrase gene (lsch) from this bacterium was found to be composed of 1,275 Up coding for a protein of 424 amino acids, with an estimated molecular weight of 47 kDa. The bacterial levansucrase gene was expressed in Escherichia coli DH5${\alpha}$ by its own promoter and lac promoter. The recombinant levansucrase was produced in soluble form with 170U of levansucrase activity from 1-ml E. coii culture broth. The expressed enzyme from the clone showed similar biochemical properties, such as size of active levansucrase, degree of branching, and optimum temperature, with P.aurantiaca 5-4380 levansucrase.

• Korean Journal of Microbiology
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• v.40 no.4
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• pp.342-347
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• 2004

A bacterial strain YC4963 with antifungal activity against Colletotrichum orbiculare, a causal organism of cucumber anthracnose was isolated from the rhizosphere soil of Siegesbeckia pubescens Makino in Korea. Based on physiological and biochemical characteristics and 16S ribosomal DNA sequence analysis, the bac­terial strain was identified as Pseudomonas aurantiaca. The bacteria also inhibited mycelial growth of several plant fungal pathogens such as Botrytis cinerea, Fusarium oxysporum and Rhizoctonia solani on PDA and 0.1 TSA media. The antifungal activity was found from the culture filtrate of this isolate and the active compound was quantitatively bound to XAD adsorption resin. The antibiotic compound was purified and identified as phenazine-l-carboxylic acid on the basis of combined spectral and chemical analyses data. This is the first report on the production of phenazine-l-carboxylic acid by Pseudomonas aurantiaca.

• Journal of Microbiology and Biotechnology
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• v.19 no.12
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• pp.1688-1694
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• 2009

A novel strain of fluorescent pseudomonad (PB-St2) was isolated from surface-sterilized stems of sugarcane grown in Pakistan. The bacterium was identified as Pseudomonas aurantiaca on the basis of 16S rRNA gene sequence analysis and results from physiological and biochemical characteristics carried out with API50 CH and QTS 24 bacterial identification kits. Assays using substrate-specific media for enzymes revealed lipase and protease activities but cellulase, chitinase, or pectinase were not detected. The bacterium was unable to solubilize phosphate or produce indole acetic acid. However, it did produce HCN, siderophores, and homoserine lactones. In dual culture assays on agar, the bacterium showed antifungal activity against an important pathogen of sugarcane in Pakistan, namely Colletotrichum falcatum, as well as for pathogenic isolates of Fusarium oxysporium and F. lateritium but not against F. solani. The antifungal metabolites were identified using thin-layer chromatography, UV spectra, and MALDI-TOFF spectra and shown to be phenazine-1-carboxylic acid (PCA), 2-hydroxyphenazine (2-OH-PHZ), and N-hexanoyl homoserine lactone (HHL) (assessed using only TLC data). The capacity of this bacterium to produce HCN and 2-OH-PHZ, as well as to inhibit the growth of C. falcatum, has not been previously reported.

• Proceedings of the Korean Magnetic Resonance Society Conference
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• 2002.08a
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• pp.79-79
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• 2002

• Research in Plant Disease
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• v.16 no.2
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• pp.158-162
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• 2010

Four promising biocontrol agents against Phytophthora capsici were selected from 507 bacterial isolates collected from rhizosphere soils and roots of pepper plants. In vitro experiment, these four biocontrol agents inhibited mycelial growth, germination of cystospores, and formation of zoosporangia and zoospores of Phytophthora capsici. In the pot experiment, the four biocontrol agents showed control efficiency higher than 70%. In greenhouse experiment, the isolates G28-6 gave the control value of 79.4%. These four biocontrol agents successfully colonized in the population density beyond 105 cfu/g on roots of pepper in vitro. The isolates G28-6 was identified as Pseudomonas aurantiaca, based on its cultural, morphological, and biochemical characterization and 16S rRNA gene sequence analysis.