• Journal of Microbiology and Biotechnology
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• 제16권12호
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• pp.1995-1999
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• 2006

Pseudomonas sp. K82 cultured in p-hydroxybenzoate induces protocatechuate 4,5-dioxygenase (PCD 4,5) for p-hydroxybenzoate degradation. In this study, a 6.0-kbp EcoR1 fragment containing p-hydroxybenzoate degradation genes was cloned from the genome of Pseudomonas sp. K82. Sequence analysis identified four genes, namely, pcaD, pcaA, pcaB, and pcaC genes known to be involved in p-hydroxybenzoate degradation. Two putative 4-hydroxyphenylpyruvate dioxygenases and one putative oxidoreductase were closely located by the p-hydroxybenzoate degradation genes. The gene arrangement and sequences of these p-hydroxybenzoate degradation genes were similar to those of Comamonas testosteroni and Pseudomonas ochraceae. PcaAB (PCD4,5) was overexpressed in the expression vector pGEX-4T-3, purified using a GST column, and confirmed to have protocatechuate 4,5-dioxygenase activity. The N-terminal amino acid sequences of overexpressed PCD4,5 were identical with those of purified PCD4,5 from Pseudomonas sp. K82.

• 한국식품저장유통학회지
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• 제21권4호
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• pp.529-535
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• 2014

본 연구는 전통적으로 방부 또는 살균효과가 있어 떡 등에 첨가되거나 저장 및 포장에 이용되었던 자생식물 15종 21건에 대하여 데하이드로초산(sodium dehydroacetate), 소르빈산(sorbic acid), 안식향산(benzoic acid), 파라옥시안 식향산메틸(methyl p-hydroxybenzoate), 파라옥시안식향산에틸(ethyl p-hydroxybenzoate), 파라옥시안식향산프로필(propyl p-hydroxybenzoate), 파라옥시안식향산이소프로필(isopropyl p-hydroxybenzoate), 파라옥시안식향산이소부틸(isobutyl p-hydroxybenzoate), 파라옥시안식향산부틸(butyl p-hydroxybenzoate), 프로피온산(propionic acid)의 함유량을 분석하였다. 솔잎, 망개잎, 대나무잎 등 15종에 대한 천연유래 보존료 함유량을 조사한 결과 솔잎, 대나무잎, 칡잎, 모시잎, 쑥, 잣나무잎 등 6종에서 안식향산, 소르빈산, 프로피온산을 확인할 수 있었다. 안식향산은 대나무잎, 모시잎, 솔잎, 쑥, 칡잎, 잣나무잎에서 8.201~21.839 mg/kg, 소르빈산은 대나무잎과 칡잎에서 5.630~24.995 mg/kg, 프로피온산은 모시잎에서 61.324~62.726 mg/kg이 검출되었다. 조사 식물 중 망개잎, 토란잎, 조릿대, 연잎, 원추리, 구절초, 떡갈나무잎, 산초잎, 감잎 등 9종에서는 천연보존성분이 검출되지 않았다.

• Journal of Microbiology
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• 제42권2호
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• pp.152-155
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• 2004

Pseudomonas sp. K82 has been reported to be an aniline-assimilating soil bacterium. However, this strain can use not only aniline as a sole carbon and energy source, but can also utilize benzoate, p-hydroxybenzoate, and aniline analogues. The strain accomplishes this metabolic diversity by using dif-ferent aerobic pathways. Pseudomonas sp. K82, when cultured in p-hydroxybenzoate, showed extradiol cleavage activity of protocatechuate. In accordance with those findings, our study attempted the puri-fication of protocatechuate 4,5-dioxygenase (PCD 4,5). However the purified PCD 4,5 was found to be very unstable during purification. After Q-sepharose chromatography was performed, the crude enzyme activity was augmented by a factor of approximately 4.7. From the Q-sepharose fraction which exhibited PCD 4,5 activity, two subunits of PCD4,5 (${\alpha}$ subunit and ${\beta}$ subunit) were identified using the N-terminal amino acid sequences of 15 amino acid residues. These subunits were found to have more than 90％ sequence homology with PmdA and PmdB of Comamonas testosteroni. The molecular weight of the native enzyme was estimated to be approximately 54 kDa, suggesting that PCD4,5 exists as a het-erodimer (${\alpha}$$_1$${\beta}$$_1$). PCD 4,5 exhibits stringent substrate specificity for protocatechuate and its optimal activity occurs at pH 9 and 15 $^{\circ}C$. PCR amplification of these two subunits of PCD4,5 revealed that the ${\alpha}$ subunit and ${\beta}$ subunit occurred in tandem. Our results suggest that Pseudomonas sp. K82 induced PCD 4,5 for the purpose of p-hydroxybenzoate degradation.

• 한국식품과학회지
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• 제4권1호
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• pp.24-28
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• 1972

Synthetic food preservatives were analysed in foods collected in Seoul area on Aug. 10, 1971. Sorbic acid, benzoic acid, dehydroacetic acid and butyl p-hydroxybenzoate were determined by the simultaneous gas chromatography using FID at $200{\circ}C$ and a column of Chromosorb W coated with 5% $DGS{\sim}1%\;H_3PO_4$. The recovery rates of each preservative were from 76.7% to 96.3%. The calibration curves show linearity within a range from 0.3 to $2.5{\mu}g$ of standard preservatives. The results obtained were as follows: 1) Benzoic acid was used as well as butyl p-hydroxybenzoate in soy. 2) Sorbic acid was not found in soy. 3) From all breads and biscuits benzoic acid was found as trace. 4) Detected preservatives were below the range of permitted limit. 5) From 2 soy among 15 samples dehydroacetic acid was found.

• Journal of Microbiology
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• 제44권6호
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• pp.632-640
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• 2006

In this study, the biodegradative activities of monocyclic aromatic compounds were determined from the multi-drug resistant (MDR) Acinetobacter baumannii, which were studied in the form of clinical isolates from a hospital in Korea. These bacteria were capable of biodegrading monocyclic aromatic compounds, such as benzoate and p-hydroxybenzoate. In order to determine which pathways are available for biodegradation in these stains, we conducted proteome analyses of benzoate, and p-hydroxybenzoate-cultured A. baumannii DU202, using 2-DE/MS analysis. As genome DB of A. baumannii was not yet available, MS/MS analysis or de novo sequencing methods were employed in the identification of induced proteins. Benzoate branch enzymes [catechol 1,2-dioxygenase (CatA) and benzoate dioxygenase $\alpha$ subunit (BenA)] of the $\beta$-ketoadipate pathway were identified under benzoate culture condition and p-hydroxybenzoate branch enzymes [protocatechuate 3,4-dioxygenas $\alpha$ subunit (PcaG) and 3-carboxy-cis,cis-muconate cycloisomerase (PcaR)] of the $\beta$-ketoadipate pathway were identified under p-hydroxybenzoate culture condition, respectively, thereby suggesting that strain DU202 utilized the $\beta$-ketoadipate pathway for the biodegradation of monocyclic aromatic compounds. The sequence analysis of two purified dioxygenases (CatA and PcaGH) indicated that CatA is closely associated with the CatA of Acinetobacter radiresistance, but PcaGH is only moderately associated with the PcaGH of Acinetobacter sp. ADPI. Interestingly, the fused form of PcaD and PcaC, carboxymuconolactone decarboxylase (PcaCD), was detected on benzoate-cultured A. baumannii DU202. These results indicate that A. baumannii DU202 exploits a different $\beta$-ketoadipate pathway from other Acinetobacter species.

• KSBB Journal
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• 제19권1호
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• pp.50-56
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• 2004

본 연구의 목적은 방향족 화한물인 aniline, benzoate, p-hydroxybenzoate를 분해할 수 있는 Delftia sp. JK-2에서 이들 각 기질에서 배양시 다른 종류의 dioxygenases를 분리 정제하고, 정제된 dioxygenases의 특성을 조사하기 위하여 실시하기 위한 것이다. 기질로서 benzoate, aniline, 또는 p-hydroxybenzoate에 따라 분리된 dioxygenases는 각각 catechol 1,2-dioxygenase (C1 ,2O), catechol 2,3-dioxygenase(C2, 3O), 그리고 protocatechuate 4,5-dioxygenase (4,5-PCD)였다. 각 dioxygenases의 특성을 조사하기 위하여 먼저 benzoate, aniline 또는 p-hydroxybenzoate에서 배양한 Delftia sp. JK-2 세포를 초음파 분쇄기로 파쇄하여, ammonium sulfate precipitation, DEAE-sepharose, 그리고 Q-sepharose의 순서로 정제하여 농축하였다. 정제$.$농축된 dioxygenases의 특이 활성도를 보면 C1, 2O는 3.3 unit/mg, C2, 3O는 4.7unit/mg이고, 4,5-PCD는 2.0 unit/mg이다 C1, 2O와 C2, 3O의 기질 특이성 조사에서는 catechol과 4-methylcatechol에서 두 효소 모두 효소 활성이 나타났으며, C1. 2O에서는 3-methylcatechol에서 약간의 활성이 확인되었고, 4,5-PCD는 protocatechuate에서만 효소 활성을 보여주었다. C1l, 2O와 C2, 3O는 3$0^{\circ}C$와 pH 8.0에서 최적의 활성을 나타내는 것으로 조사되었으며, 4,5-PCD는 3$0^{\circ}C$와 pH 7.0에서 최적의 활성이 조사되었다. Delftia sp. JK-2에서 정제된 C1, 2O와 C2, 3O의 효소활성은 Ag$^{+}$, Hg$^{+}$, 그리고 Cu$^{2+}$에 의해 억제되는 것으로 나타났으며, 4,5-PCD의 경우에는 Ag$^{+}$, Hg$^{+}$, 그리고 Cu$^{2+}$ 뿐만 아니라 Fe$^{3+}$ 에 이해서도 효소 활성이 억제되는 것이 확인되었다. C1, 2O, C2, 3O, 4,5-PCD의 분자량은 SDS-PAGE에 의해 각각 60kDa, 35kDa, 62kDa로 측정되었다.

• 대한화학회지
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• 제18권1호
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• pp.72-75
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• 1974

• Journal of Microbiology and Biotechnology
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• 제28권3호
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• pp.439-447
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• 2018

The aromatic compound p-hydroxybenzoate (PHBA) is an important material with multiple applications, including as a building block of liquid crystal polymers in chemical industries. The cytochrome P450 (CYP) enzymes are beneficial monooxygenases for the synthesis of chemicals, and CYP53A15 from fungus Cochliobolus lunatus is capable of executing the hydroxylation from benzoate to PHBA. Here, we constructed a system for the bioconversion of benzoate to PHBA in Escherichia coli cells coexpressing CYP53A15 and human NADPH-P450 oxidoreductase (CPR) genes as a redox partner. For suitable coexpression of CYP53A15 and CPR, we originally constructed five plasmids in which we replaced the N-terminal transmembrane region of CYP53A15 with a portion of the N-terminus of various mammalian P450s. PHBA productivity was the greatest when CYP53A15 expression was induced at $20^{\circ}C$ in $2{\times}YT$ medium in host E. coli strain ${\Delta}gcvR$ transformed with an N-terminal transmembrane region of rabbit CYP2C3. By optimizing each reaction condition (reaction temperature, substrate concentration, reaction time, and E. coli cell concentration), we achieved 90% whole-cell conversion of benzoate. Our data demonstrate that the described novel E. coli bioconversion system is a more efficient tool for PHBA production from benzoate than the previously described yeast system.

• 대한화장품학회지
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• 제8권1호
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• pp.75-83
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• 1980

• 대한화장품학회지
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• 제11권1호
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• pp.13-20
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• 1985

Polyoxyethylene계 유화제를 이용한 유화계에 있어서 talc 및 $TiO_2$에 의한 methyl p-hydroxy benzoate의 흡착과 그에 따른 방부력 감소를 여러 가지 분석방법과 미생물 실험을 통하여 측정하였다. 이 유화계에 있어서 methyl p-hydroxy benzoate는 talc 및 $TiO_2$에 흡착되어 방부력이 감소함을 알 수 있으며 방부력 저하에 미치는 영향은 $TiO_2$보다 talc가 더 크고, talc에 의한 methyl p-hydroxy benzoate의 흡착량은 $20^{\circ}C$에서 다음과 같은 식으로 나타낼 수 있었다. 즉 $a=11.511C^{0.747}$