• Journal of Microbiology and Biotechnology
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• 제12권1호
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• pp.31-38
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• 2002

Bacteria have evolved various types of resistance mechanism to toxic heavy metals, such as arsenic and antimony. An arsenical and antimonial resistant bacterium was isolated from a shallow creek draining a coal-mining area near Taebaek City, in Kangwon-Do, Korea. The isolated bacterium was identified and named as Pseudomonas sp. KM20 after biochemical and physiological studies were conducted. A plasmid was identified and its function was studied. Original cells harboring the plasmid were able to grow in the presence of 15 mM sodium arsenite, while the plasmid-cured (plasmidless) strain was sensitive to as little as 0.5 mM sodium arsenate. These results indicated that the plasmid of Pseudomonas sp. KM20 does indeed encode the arsenic resistance determinant. In growth experiments, prior exposure to 0.1 mM arsenate allowed immediate growth when they were challenged with 5 mM arsenate, 5 mM arsenite, or 0.1 mM antimonite. These results suggested that the arsenate, arsenite, and antimonite resistance determinants of Pseudomonas sp. KM20 plasmid were indeed inducible. When induced, plasmid-bearing resistance cells showed a decreased accumulation $of\;73^As$ and showed an enhanced efflux $of\;^73As$. These results suggested that plasmid encoded a transport system that extruded the toxic metalloids, resulting in the lowering of the intracellular concentration of toxic oxyanion. In a Southern blot study, hybridization with an E. coli R773 arsA-specific probe strongly suggested the absence of an arsA cistron in the plasmid-associated arsenical and antimonial resistance determinant of Pseudomonas sp. KM20.

• Journal of Microbiology and Biotechnology
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• 제13권6호
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• pp.1001-1007
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• 2003

Metal ions contamination may inhibit microorganisms involved in the biodegradation of organic compounds and affect biodegradation rates. Therefore, it is likely that bioremediation of xenobiotics-contaminated soils and waste will require inoculation with efficient biodegrading microbial communities, with capabilities of being resistant to heavy metals as well. Two different transconjugants (Pseudomonas sp. KMl2TC and P. aeruginosa TC) were constructed by conjugation experiments. Results on MIC, induction and growth inhibition strongly indicated that arsenic-resistant plasmid, pKM20, could be mobilized, and the newly acquired phenotype of pKM20 was not only expressed but also well regulated, resulting in newly acquired resistances to $As^{5+},\;As^{3+},\;and\;Sb^{3+} in\;addition\;to\;Cd^{2+},\;Zn^{2+},\;and\;Hg^{2+}$. The phenol- degradation efficiencies of Pseudomonas sp. KMl2TC were maintained significantly even at high heavy metal concentrations at which these efficiencies of P. aeruginosa TC were completely impaired. The results in this study on the effects of heavy metals on phenol degradation, especially after conjugation, are the first ever reported. All the results described in this study encourage to establish a goal of making "designer biocatalysts" which could degrade certain xenobiotics in the area contaminated with multiple heavy metals.

• 한국수산과학회지
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• 제29권5호
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• pp.637-642
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• 1996

해양으로부터 alginate lyase분해능이 있는 Pseudomonas sp.을 분리하여 이 균으로부터 alginate lyase 유전자를 E. coli에 cloning 하여 재조합 alginate lyase의 특성을 검토하였다. 재조합 alginate lyase는 E. coli에서 $50\%$ 이상이 expression되었다. 또한 생산된 재조합 alginate lyase는 세포외로의 분비가 없었으며, 전자현미경으로 확인한 결과 대부분이 inclusion body를 형성하지 않고 세포질내에서 soluble한 형태로 존재하고 있는 것으로 나타났다. 활성을 나타내는 최적 조건은 온도 $20^{\circ}C$, pH 7.0으로 보여주고 있으며 기질에 대한 Km값은 $0.4\%$, Vmax는 625 unit/mg이었다.

• 한국미생물·생명공학회지
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• 제24권3호
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• pp.282-289
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• 1996

2,3-DHBP dioxygenase was purified from E. coli CK1092 carrying the pcbC gene, which was cloned from 4-chlorobiphenyl-degrading Pseudomonas sp. P20. Purification of this enzyme was done by acetone precipitation, DEAE- Sephadex A-25 ion exchange chromatography, and preparative gel electrophoresis. The molecular weight of subunit was 34 kDa determined by SDS-PAGE, and that of native enzyme was about 270 kDa. It suggests that this enzyme consist of eight identical subunits. This enzyme was specifically active against only 2,3-DHBP as a substrate with 18 $\mu$M of Km value, but not catechol, 3-methylcatechol, 4-methylcatechol and 4-chlorocatechol. The optimal pH and temperature of 2,3-DHBP dioxygenase were pH 8.0 and 40-60$\circ$C. The enzyme was inhibited by Cu$^{2+}$, Fe$^{2+}$ and Fe$^{3+}$ ions, and was inactivated by H$_{2}$0$_{2}$2 and EDTA. The lower concentrations of some organic solvents such as acetone and ethanol don't stabilize the activity of 2,3-DHBP dioxygenase. The enzyme was completely inactivated by adding the reagents such as N-bromosuccinimide, iodine and p- diazobenzene sulfonic acid.

• Environmental Engineering Research
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• 제15권2호
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• pp.57-62
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• 2010

The purpose of this study was to investigate the effect of mixing methods on the biodegradation of sorbed naphthalene and phenanthrene in soils. Biodegradation was initiated by inoculating Pseudomonas sp. KM1 into equilibrated soil slurry vials. Four different mixing methods, including no mixing, orbital shaking, rolling and rotating were utilized to enhance the biodegradation of both naphthalene and phenanthrene. The experimental results showed that the sorbed compounds were more effectively biodegraded with rolling and rotating mixing methods. The sorbed naphthalene concentrations were reduced to 0 mg/kg via the rolling and rotating methods. However, with no mixing and the orbital shaking methods, the sorbed naphthalene concentrations were comparatively high, ranging from 2.59 to 20.45 mg/kg. Similar trends were observed for the biodegradation of phenanthrene, but the concentrations remaining were higher than those of naphthalene, due to the limited bioavailability of the sorbed phenanthrene. The rolling and rotating mixing methods are suggested can distribute bacteria uniformly in the slurry system; improve the mass transfer rate and the probability of physical contact between bacteria and the sorbed contaminants, resulting in higher bioavailability of the contaminants.