• Journal of Microbiology and Biotechnology
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• 제13권3호
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• pp.463-468
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• 2003

Polychlorinated biphenyl are toxic pollutants and their degradation is quite slow in the environment. Recently, interest if bioremediation using PCB-degrading bacteria has increaset,. In a previous report, plant terpenes (p-cymene, (S)-(-)-limonene, ${\alpha}-pynene$, and ${\alpha}-terpinene$) have been found to be utilized by a PCB degrader and to induce the biphenyl dioxygenase gene in pure culture. In this study, Pseudomonas pseudoalcaligenes KF707, a PCB-degrading Gram-negative soil bacterium, was used to determine whether the terpene stimulation of PCB degrader occurred in the natural environment. First, P. pseudoalcaligenes KF707 was genetically tagged using a transposon with gfp (green fluorescent protein) as a reporter gone. The population dynamics of P. pseudoalcaligenes KF707 harboring gfp gene in a PCB-contaminated environment was examined with or without terpenoids added to the microcosm. About 10-100-fold increase was found in the population of PCB degraders when terpene was added, compared with control (non-terpenes samples and biphenyl added samples). It was proposed that the gfp-monitoring system is very useful and terpenes enhance the survivability of PCB degraders in PCB-contaminated environments.

• 한국미생물·생명공학회지
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• 제22권1호
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• pp.37-44
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• 1994

The homology among the genes coding for degradation of bipheny(BP) and 4-chlorobiphenyl(4CB) was comparatively analyzed by Southern hybridization in several BP/4CB degrading bacterial strains. As the hybridization results of their genomic DNAs with pcbABCD as the DNA probe, the group of Pseudomonas sp. DJ-12. P08 and P27 strain was separated by the group of P20 and P1242 strains. The P. pseudoalcaligenes KF707 showed the hybidization signal which was homologous to the group of DJ-12, but they had different restriction endonuclease sites. The pcbAB genes in pCUl recombinant plasmid from Pseudomonas sp. DJ-12 appeared to be homologous to pchAB genes in pKTF20 cloned from P. pseudoalcaligenes KF707, but the C genes in both strains were not homologous. The bphABC in pKTF20 showed the signals homologous to the cbp ACB in pAW6194 cloned from P. putida OU83, but homologous signal was not found botween the pcbABCD genes in pCUl and the cbpADCB genes in pAW6194 recombbinant plasmid.

• 환경생물
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• 제22권1호
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• pp.213-219
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• 2004

동굴 내 정점별 세균 군집 구조를 분석하기 위하여 PCf amplified 16S rDNA denaturing gradient gel electrophoresis(DGGE)를 적용하였다. DGGE는 동일한 분자량을 갖는 dsDNA band라고 할지라도, 각각의 염기서열 차이에 따라 전기영동 상에서 고유한 band양상을 나타낼 수 있다. eubacteria의 16S rDNA V3region을 증폭하기 위해 GC341F와 PRUN518r을 primer로 사용하여 지하수내에 미생물 군집의 다양성과 유사성을 분석하였다. DGGE band 양상을 통해 동굴내의 세균 군집 구조는 외부 환경에 비해 상대적으로 종다양성이 낮으며 동굴내 에서 특이적으로 서식하는 종이 있음을 확인하였다. 또한 유기 영양물질의 공급이 제한되어 있는 동굴에서 구아노가 주요 유기 영양물질의 공급원으로서 큰 영향을 미치고 있는 것으로 파악되었다. DGGE 상의 일부 band의 염기서열분석 결과 Pseudomonas sp. NZ060과 Pseudomonas pseudoalcaligenes, uncultured Variovorax sp., soil bacterium NS7로 동정되었다.

• 생명과학회지
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• 제25권5호
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• pp.487-495
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• 2015

Pseudomonas pseudoalcaligenes KF707에서 정제한 protocatechuate 3,4-dioxygenase의 특징을 조사하기 위하여 pH안정성, 화학적 저해, 화학적 수식과 pH의존성 반응 상수에 대한 실험을 수행하였다. 이 효소는 pH 4.5~10.7에서 안정하였다. L-ascorbate와 glutathione은 Kis가 각각 0.17 mM과 0.86 mM인 경쟁적 저해제였으며, DL-dithiothreitol은 Kis 1.57 mM 및 Kii 8.08 mM의 비경쟁적 저해패턴을 나타내었다. Potassium cyanide, p-hydroxybenzoate 및 sodium azide는 Kis가 각각 55.7 mM, 0.22 mM 및15.64 mM이었으며, Kii는 각각94.1 mM, 8.08 mM, 및 662.64 mM인 비경쟁적 저해패턴을 나타내었다. $FeCl_{2}$는 Kis가 $29{\mu}M$로 가장 우수한 경쟁적 저해제였으며, $FeCl_{3}$, $MnCl_{2}$, $CoCl_{2}$, $HgCl_{2}$, $AlCl_{3}$도 각각 Kis가 1.21 mM, 0.85 mM, 3.98 mM, 0.17 mM 및 0.21 mM인 경쟁적 저해패턴을 보였다. 한편, 다른 금속이온들은 비경쟁적 저해패턴을 나타내었다. pH의존성 반응상수의 실험결과로부터 pK 6.2와 9.4의 촉매부위와 pK 5.5와 9.0의 결합부위가 존재함을 알 수 있었다. Lysine, cysteine, tyrosine, carboxyl과 histidine은 각각의 고유한 화학적 수식제에 의해 수식되었는데, 이는 이들 잔기들이 결합과 촉매에 관여한다는 것을 나타낸다. 위 결과를 토대로 화학적 메커니즘을 제시한다.

• 미생물학회지
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• 제38권4호
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• pp.275-275
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• 2002

Pseudomonas sp. S-47 is capable of catabolizing 4-chlorobenzoate (4CBA) as carbon and energy sources under aerobic conditions via the mesa-cleavage pathway. 4CBA-dioxygenase and 4CBA-dihydrodiol dehydrogenase (4CBA-DD) catalyzed the degradation af 4CBA to produce 4-chlorocatechol in the pathway. In this study, the xylL gene encoding 4CBA-DD was cloned from the chromosomal DNA of Pseudomonas sp. S-47 and its nucleotide sequence was analyzed. The xylL gene was found to be composed of 777 nucleotide pairs and to encode a polypeptide of 28 kDa with 258 amino acid residues. The deduced amino acid sequence of the dehydrogenase (XylL) from strain S-47 exhibited 98% and 60% homologies with these of the corresponding enzymes, Pseudomonas putida mt-2 (XyIL) and Acinetobacter calcoaceticus (BenD), respectively. However, the amino arid sequences show 30% or less homology with those of Pseudomonas putida (BnzE), Pseudomonas putida Fl (TodD), Pseudomonas pseudoalcaligenes KF707 (BphB), and Pseudomonas sp. C18 (NahB). Therefore, the 4CBA-dihydrodiol dehdrogenase of strain S-47 belongs to the group I dehydrogenase involved in the degradation of mono-aryls with a carboxyl group.

• Journal of Microbiology
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• 제38권4호
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• pp.275-280
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• 2000

Pseudomonas sp. S-47 is capable of catabolizing 4-chlorobenzoate (4CBA) as rarbon and energy sources under aerobic conditions via the mesa-cleavage pathway. 4CBA-dioxygenase and 4CBA-dihydrodiol dehydrogenase (4CBA-DD) catalyzed the degradation af 4CBA to produce 4-chlorocatechol in the pathway. In this study, the xylL gene encoding 4CBA-DD was cloned from the chromosomal DNA of Pseudomonas sp. S-47 and its nucleotide sequence was analyzed. The xylL gene was found to be composed of 777 nucleotide pairs and to encode a polypeptide of 28 kDa with 258 amino acid residues. The deduced amino acid sequence of the dehydrogenase (XylL) from strain S-47 exhibited 98% and 60% homologies with these of the corresponding enzymes, Pseudomonas putida mt-2 (XyIL) and Acinetobacter calcoaceticus (BenD), respectively. However, the amino arid sequences show 30% or less homology with those of Pseudomonas putida (BnzE), Pseudomonas putida Fl (TodD), Pseudomonas pseudoalcaligenes KF707 (BphB), and Pseudomonas sp. C18 (NahB). Therefore, the 4CBA-dihydrodiol dehdrogenase of strain S-47 belongs to the group I dehydrogenase involved in the degradation of mono-aryls with a carboxyl group.

• Journal of Microbiology and Biotechnology
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• 제10권2호
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• pp.258-263
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• 2000

2-Hydroxy-6-oxo-6phenylhexa-2,4-dienoate (HOPDA) hydrolase catalyzes the hydrolytic cleavage of HOPDA to bemzpate and 2-hydroxypenta-2, 4-dienoate (HPD) during microbial catabolism of biphenyl and polychlorinated biphenyls. A HOPDA hydrolase gene (pcbD) was isolated from the genomic library of Pseudomonas sp. P20 and designated as pCNUO1201; a 7.5-kb XbaI DNA fragment from Pseudomonas sp. P20 was inserted into the pBluescript SK(+) XbaI site. E. coli HB101 harboring pCNU1201 exhibited HOPDA hydrolase activity. The open reading frame (ORF) corresponding to the pcbD gene consisted of 855 base pairs with an ATG initiation codon and a TGA termination codon. The ORF was preceded by a rebosome-binding sequence of 5'-TGGAGC-3' and its G+C content was 55 mol%. The pcbD gene of Pseudomonas sp. P20 was located immedeately downstream of the pcbC gene encoding 2,3- dihydroxybiphenyl 1,2-dioxygenase, and approximately 4-kb upstream of the pcbE gene encoding HPD hydratase. The pcbK gene was able to encode a polypeptide with a molecular weight of 31,732 containing 284 amino acid residues. The deduced amino acid sequence of the HOPDA hydrolase of Pseudomonas sp. P20 exhibited high identity (62%) with those of the HOPDA hydrolases of P. putida KF715, P. pseudoalcaligenes KF707, and Burkholderia cepacia LB400, and also significant homology with those of other hydrolytic enzymes including esterase, transferase, and peptidase.

• Journal of Microbiology
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• 제38권1호
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• pp.31-35
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• 2000

A suitable packing material for biofiltration of monoaromatic solvent vapors was selected among various types of packing materials such as peat, bark chips, vermiculite, and Hydroballs. A previously isolated strain, Pseudomonas pseudoalcaligenes BTXO2, which could utilize toluene, m-and p-xylene as carbon and energy sources was used as a biofilter inoculum. Four glass biofilters (6 cm dia. x 60 cm) were individually packed with each of the packing materials and solvent vapors were passed through the columns. During three weeks of peat biofilter operation, average removal efficiencies of toluene, m-and p-xylene were 90.4%, 95.3%, and 82.1%, respectively. With the other packings, the efficiencies were in the range of 10.1 to 58.6％ which were significantly lower than those of the peat biofilter. The peat biofilter was continually operated for approximately nine months and the biofilter sustained its degradation activity during the operation period with minimal maintenance. At steady state, average removal rates of toluene, m- and p-xylene vapors were estimated as 14.2, 5.5, and 8.1 g m$\^$-3/ packing h$\^$-1/, respectively.

• 한국균학회지
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• 제22권1호
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• pp.50-61
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• 1994

토양으로부터 분리한 380개체의 균주 중 Fusarium solani에 대해 길항능력이 있는 방선균 및 세균 42개 균주로부터 길항작용이 큰 균주로 CHA 1과 5-PFHR 6을 공시하여 동정한 결과 각각 Promicromonospora sp.와 Pseudomonas pseudoalcaligenes로 동정되었다. PDA 평판배지상에서 두 균주는 모두 F. solani의 저해작용으로 균사의 길이 생장억제, 비정상적인 균사의 분지, 세포벽 분해에 의한 돌기형성 등이 관찰되었다. F. solani 포자발아에 미치는 길항균주의 배양여과액의 작용으로 감자즙액 배지나 nutrient broth에 배양한 여과액은 포자발아 억제력이 높아 14.3%의 낮은 발아율을 보였으며, 토양침출액 단독 또는 두 영양배지에 토양침출액을 첨가하여 준비한 배양 여과액에서의 발아율은 85% 이상으로 포자발아가 촉진되었으나 S-PFHR 6의 경우 두 영양배지에 소량의 토양침출액을 첨가한 배앙여과액에서 4%의 발아율을 보였다. 자연토양에서 대형포자는 19.4%, 후막포자는 17.7%가 발아하였으나 증기살균 토양에서의 발아율은 79.7% 이상으로 증가하였다. 자연토양에 공시길항균 CHA 1과 S-PFHR 6의 균체를 처리하면 토양 정균력이 증가되어 F. solani의 대형 포자 발아율은 각 처리구에서 각각 14.7%, 11.7%로 감소하였다. 길항균을 처리한 토양에 glucose와 asparagine을 처리하면 토양의 정균력이 점차 해소되어 대형포자의 발아율은 48.0% 이상으로 증가하였다. 공시균주의 2차 대사산물에 의한 길항작용으로 두 균주의 배양여과액을 토양에 처리한 경우 CHA 1의 배앙여과액을 처리한 토양에서의 F. solani 대형포자의 발아율은 9.3%, 5-PFHR 6 처리구에서의 발아율은 38.0%가 되었다. F. solani의 후막포자는 공시길한균주의 균체나 배양여과액 처리구에서의 발아율이 자연토양에서의 발아율보다 낮았으며 이러한 현상은 영앙원을 첨가해 주어도 변하지 않았다.