Biomedical subjects

Kil Yong Kim

Publications and source records attributed to Kil Yong Kim.

8 recordsLinked to original sources

PubMed2006

Rhizobium etli USDA9032 engineered to produce a phenazine antibiotic inhibits the growth of fungal pathogens but is impaired in symbiotic performance.

Phenazine production was engineered in Rhizobium etli USDA9032 by the introduction of the phz locus of Pseudomonas chlororaphis O6. Phenazine-producing R. etli was able to inhibit the growth of Botrytis cinerea and Fusarium oxysporum in vitro. Black bean inoculated with phenazine-producing R. etli produced brownish Fix(-) nodules.

Anti-Bacterial Agents↗

PubMed2006

GacS-dependent production of 2R, 3R-butanediol by Pseudomonas chlororaphis O6 is a major determinant for eliciting systemic resistance against Erwinia carotovora but not against Pseudomonas syringae pv. tabaci in tobacco.

Root colonization by a plant-beneficial rhizobacterium, Pseudomonas chlororaphis O6, induces disease resistance in tobacco against leaf pathogens Erwinia carotovora subsp. carotovora SCC1, causing soft-rot, and Pseudomonas syringae pv. tabaci, causing wildfire. In order to identify the bacterial determinants involved in induced systemic resistance against plant diseases, extracellular components produced by the bacterium were fractionated and purified. Factors in the culture filtrate inducing systemic resistance were retained in the aqueous fraction rather than being partitioned into ethyl acetate. Fractionation on high-performance liquid chromatography followed by nuclear magnetic resonance mass spectrometry analysis identified the active compound as 2R, 3R-butanediol. 2R, 3R butanediol induced systemic resistance in tobacco to E. carotovora subsp. carotovora SCC1, but not to P. syringae pv. tabaci. Treatment of tobacco with the volatile 2R, 3R-butanediol enhanced aerial growth, a phenomenon also seen in plants colonized by P. chlororaphis O6. The isomeric form of the butanediol was important because 2S, 3S-butandiol did not affect the plant. The global sensor kinase, GacS, of P. chlororaphis O6 was a key regulator for induced systemic resistance against E. carotovora through regulation of 2R, 3R-butanediol production. This is the first report of the production of these assumed fermentation products by a pseudomonad and the role of the sensor kinase GacS in production of 2R, 3R-butanediol.

Bacterial Proteins↗

PubMed2005

Selective preparation of N-acetyl-D-glucosamine and N,N'-diacetylchitobiose from chitin using a crude enzyme preparation from Aeromonas sp.

A bacterium, Aeromonas sp. GJ-18, having strong chitinolytic activity was isolated from coastal soil and used for crude enzyme preparations. This enzyme preparation contained N-acetyl-D-glucosaminidase and N,N'-diacetylchitobiohydrolase. N-Acetyl-D-glucosaminidase was inactive above 50 degrees C, but N,N'-diacetylchitobiohydrolase was stable at this temperature. Utilizing the temperature sensitivities of the chitin degradation enzymes in crude enzyme preparation, N-acetyl-D-glucosamine (GlcNAc) and N,N'-diacetylchitobiose [(GlcNAc)(2)] were selectively produced from chitin. At 45 degrees C, GlcNAc was produced as a major hydrolytic product (94% composition) with a yield of 74% in 5 d, meanwhile at 55 degrees C (GlcNAc)(2) was the major product (86%) with a yield of 35% within 5 d.

Acetylglucosamine↗

PubMed2005

Active changes of lignification-related enzymes in pepper response to Glomus intraradices and/or Phytophthora capsici.

The activities of enzymes responsible for lignification in pepper, pre-inoculation with arbuscular mycorrhizal (AM) fungus of Glomus intraradices and/or infection with pathogenic strain of Phytophthora capsici, and the biological control effect of G. intraradices on Phytophthora blight in pepper were investigated. The experiment was carried out with four treatments: (1) plants pre-inoculated with G. intraradices (Gi), (2) plants pre-inoculated with G. intraradices and then infected with P. capsici (Gi+Pc), (3) plants infected with P. capsici (Pc), and (4) plants without any of the two microorganisms (C). Mycorrhizal colonization rate was reduced by about 10% in pathogen challenged plants. Root mortality caused by infection of P. capsici was completely eliminated by pre-inoculation with antagonistic G. intraradices. On the ninth day after pathogen infection, Peroxidase (POD) activity increased by 116.9% in Pc-treated roots but by only 21.2% in Gi+Pc-treated roots, compared with the control, respectively. Polyphenol oxidase (PPO) and Phenylalanine ammonia-lyase (PAL) activities gradually increased during the first 3 d and dramatically decreased in Pc-treated roots but slightly decreased in Gi+Pc-treated roots, respectively. On the ninth day after pathogen infection, PPO and PAL decreased by 62.8% and 73.9% in Pc-treated roots but by only 19.8% and 19.5% in Gi+Pc-treated roots, compared with the control, respectively. Three major POD isozymes (45,000, 53,000 and 114,000) were present in Pc-treated roots, while two major bands (53,000 and 114,000) and one minor band (45,000) were present in spectra of Gi+Pc-treated roots, the 45,000 POD isozyme was significantly suppressed by G. intraradices, suggesting that the 45,000 POD isozyme was induced by the pathogen infection but not induced by the antagonistic G. intraradices. A 60,000 PPO isozyme was induced in Pc-treated roots but not induced in Gi+Pc-treated roots. All these results showed the inoculation of antagonistic G. intraradices alleviates root mortality, activates changes of lignification-related enzymes and induces some of the isozymes in pepper plants infected by P. capsici. The results suggested that G. intraradices is a potentially effective protection agent against P. capsici.

Capsicum↗

PubMed2003

2-ketogluconic acid production and phosphate solubilization by Enterobacter intermedium.

Enterobacter intermedium, isolated from grass rhizosphere, exhibited a strong ability to solubilize insoluble phosphate. This bacterium oxidized glucose to gluconic acid and sequentially to 2-ketogluconic acid (2-KGA), which was identified using HPLC and GC-MS. The ability of E. intermedium to solubilize phosphate and produce 2-KGA produce in broth medium containing different components was monitored with air and without air supply. With an air supply, the production of 2-KGA markedly increased to about 110 g/l at day 10 in media containing 0.2 M gluconic acid, while it was about 65 g/l without gluconic acid addition. With an air supply, the concentration of soluble phosphate significantly decreased to 200-250 mg/l in media containing 1% CaCO3, whereas it was about 1000 mg/l without CaCO3 addition. Without an air supply, the concentration of 2-KGA and phosphate were negligible throughout the culture period.

Culture Media↗

PubMed2003

Cloning and expression of pyrroloquinoline quinone (PQQ) genes from a phosphate-solubilizing bacterium Enterobacter intermedium.

A grass rhizosphere bacterium, Enterobacter intermedium (60-2G), has a strong ability to solubilize insoluble phosphate. Certain phosphate-solubilizing bacteria secrete gluconic acid for this process. The gluconic acid is produced by direct extracellular oxidation of glucose by a glucose dehydrogenase equipped with pyrroloquinoline quinone (PQQ) as a cofactor. A pqq gene cluster producing PQQ was detected in E. intermedium and this sequence conferred phosphate-solubilizing activity to Escherichia coli DH5alpha. The 6,783-bp pqq sequence had six open reading frames (pqqA, B, C, D, E, and F) and showed 50-95% homology to pqq genes of other bacteria. E. coli DH5alpha expressing the E. intermedium pqq genes solubilized phosphate from hydroxyapatite after a pH drop to pH 4.0, which paralleled in time the secretion of gluconic acid. We speculate that production of PQQ in E. coli DH5alpha expressing the pqq cluster activates an endogenous glucose dehydrogenase to permit gluconic acid secretion that solubilizes the insoluble phosphate.

Cloning, Molecular↗

PubMed2003

Extracellular proteins involved in soybean cultivar-specific nodulation are associated with pilus-like surface appendages and exported by a type III protein secretion system in Sinorhizobium fredii USDA257.

Several gram-negative plant and animal pathogenic bacteria have evolved a type III secretion system (TTSS) to deliver effector proteins directly into the host cell cytosol. Sinorhizobium fredii USDA257, a symbiont of soybean and many other legumes, secretes proteins called Nops (nodulation outer proteins) into the extracellular environment upon flavonoid induction. Mutation analysis and the nucleotide sequence of a 31.2-kb symbiosis (sym) plasmid DNA region of USDA257 revealed the existence of a TTSS locus in this symbiotic bacterium. This locus includes rhc (rhizobia conserved) genes that encode components of a TTSS and proteins that are secreted into the environment (Nops). The genomic organization of the TTSS locus of USDA257 is remarkably similar to that of another broad-host range symbiont, Rhizobium sp. strain NGR234. Flavonoids that activate the transcription of the nod genes of USDA257 also stimulate the production of novel filamentous appendages known as pili. Electron microscope examination of isolated pili reveals needle-like filaments of 6 to 8 nm in diameter. The production of the pili is dependent on a functional nodD1 and the presence of a nod gene-inducing compound. Mutations in several of the TTSS genes negate the ability of USDA257 to elaborate pili. Western blot analysis using antibodies raised against purified NopX, Nop38, and Nop7 reveals that these proteins were associated with the pili. Mutations in rhcN, rhcJ, rhcC, and ttsI alter the ability of USDA257 to form nodules on Glycine max and Macroptilium atropurpureum.

Bacterial Proteins↗

PubMed2003

Citrate synthase mutants of Sinorhizobium fredii USDA257 form ineffective nodules with aberrant ultrastructure.

The tricarboxylic acid (TCA) cycle plays an important role in generating the energy required by bacteroids to fix atmospheric nitrogen. Citrate synthase is the first enzyme that controls the entry of carbon into the TCA cycle. We cloned and determined the nucleotide sequence of the gltA gene that encodes citrate synthase in Sinorhizobium fredii USDA257, a symbiont of soybeans (Glycine max [L.] Merr.) and several other legumes. The deduced citrate synthase protein has a molecular weight of 48,198 and exhibits sequence similarity to citrate synthases from several bacterial species, including Sinorhizobium meliloti and Rhizobium tropici. Southern blot analysis revealed that the fast-growing S. fredii strains and Rhizobium sp. strain NGR234 contained a single copy of the gene located in the bacterial chromosome. S. fredii USDA257 gltA mutant HBK-CS1, which had no detectable citrate synthase activity, had diminished nodulation capacity and produced ineffective nodules on soybean. Light and electron microscopy observations revealed that the nodules initiated by HBK-CS1 contained very few bacteroids. The infected cells contained large vacuoles and prominent starch grains. Within the vacuoles, membrane structures that appeared to be reminiscent of disintegrating bacteroids were detected. The citrate synthase mutant had altered cell surface characteristics and produced three times more exopolysaccarides than the wild type produced. A plasmid carrying the USDA257 gltA gene, when introduced into HBK-CS1, was able to restore all of the defects mentioned above. Our results demonstrate that a functional citrate synthase gene of S. fredii USDA257 is essential for efficient soybean nodulation and nitrogen fixation.

Citrate (si)-Synthase↗