• 미생물학회지
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• 제29권2호
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• pp.92-96
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• 1991

The soluble cytochrome c-551 of photosynthetic bacterium, Rhodopseudomonas gelatinosa ATCC 17013 was purified through a sequene of four step chromatography including CM-cellulose ion-exchange chromatography, DEAE-Sephacel chromatography, Sephacryl s-200 gel permeation chromatography, and HPLC (SP-5PW). The molecular weight of the purified cytochrome c-551 was 14, 600 Da, and this protein shows the absorption peak at 551 nm, 522 nm, and 417 nm as the reduced form, and at 412 nm as the oxidized form. The cytochrome c-551 seems to be a substrate for the terminal oxidase in the electron transport chain.

• 미생물학회지
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• 제30권2호
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• pp.101-107
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• 1992

화학 영양성으로 배양한 Rps. gelatinosa 에서 2회의 시토크롬 c 친화성 크로마토그래피와 DEAE-Sephacel 이온 교환 크로마토그래피 등 3 단계의 크로마토그래피를 수행하여 시토로콤 c 산화효소를 정제하였다. 정제된 시토크롬 c 산화효소는 Sephacryl S-300 에 의한 분자걍이 약 110,000 Da 이고 SDS-gel 전기영동에 의한 분자량이 약 52.000 Da 으로써 이량체일 것으로 보인다. 전제된 시토크롬 c 산화효소는 온도데 매우 불안정하고 말 심장 시토크롬 c 를 기질로 사용했을때 Km 값은 $20\mu$M, Vmax 값은 44unit/mg prot. 이며 pH 6.4 의 효소방응 최적 pH 와 25.deg.C 의 최적 온도를 보였다. 환원된 시토크롬 c 산화효소는 554, 523, 421 nm 에서 .alpha., .betha. soret 흡수대를 보였고 chromatophore 에서와 마찬가지로 KCN 과 $NaN_{3}$ 에 의해서는 효소 활성도가 저해를 받았지만 CO 와 antimycin A, myxothiazol 에 의해서는 효소 활성도가 저해를 받지 않았다. 빛을 에너지원으로 배양하거나 또는 화학영양성으로 배양하든지 모두 시토크롬 c-551 이 생성되었고 환원된 시토크롬 c-551 은 시토크롬 c 산화효소에 의해 산화되었다. 시토크롬 c-551 을 기질고 이용하였을 때 시토크롬 c 산화효소의 Km 값은 $26\mu$M 이었고 Vmax 값은 31.unit./mg prot. 로써 말심장의 시토크롬 c 를 기질로 이용할때 보다 오히려 낮았다. 이와 같은 결과로 보아 화학 영양성은 배양한 Rhodopseudomonas gelatinosa 에서 호흡에 의한 전자전달은 시토크롬 c-551 이 시토크롬 $bc_{1}$ 복합체로 부터 전자를 받아 b-형 시토크롬 c 산화효소에 전자를 전달해 주고 최정적으로 산소를 환원시킬 것으로 생각된다.

• 미생물학회지
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• 제29권6호
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• pp.380-386
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• 1991

Three types of soluble cytochrome c were purified to homogeneity from Methylobacillus sp. strain SK1 which grows only on methanol. Cytochrome c-I was purified 58.5-fold in seven steps. Cytochrome c-II and c-III were purified 57.3- and 122.1-fold in eight steps, respectively. The molecular weights of the cytochrome c-I was determined to be 12,500, while those of the cytochrome c-II and c-III were 16,000. The isoelectric points of the c-I, c-II and c-III were found to be 8.8, 6.6, and 6.6 respectively. The spectrum of reduced cytochrome c-I showed .alpha.-, .betha.-, .gamma.-peaks at 551.4, 522.2, and 416.6nm. The peaks for c-II were found at 551.0, 521.6, and 416.5nm, while those for c-III were shown at 551.2, 521.8, and 416.0 nm. The spectra of oxidized cytochrome c-I, c-II, and c-III showed .gamma.-peak at 411.8, 409.0, and 410.2 nm, respectively. The absorption coefficients of .alpha.- and .gamma.-peak for c-I in the reduced state were determined as 47 and 197 $mM^{-1}$ $cm^{-1}$ , respectively. The coefficients of .alpha.- and .gamma.-peak for c-II were determined to be 43 and 137 $mM^{-1}$ $cm^{-1}$ , while those for c-III were 41 and 172 $mM^{-1}$ $cm^{-1}$ , respectively. The c-I and c-III were found to bind carbon monoxide.

• 한국환경성돌연변이발암원학회지
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• 제26권3호
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• pp.77-85
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• 2006

2,3,7,8-Tetrachlorodibenzo-$\rho$-dioxin(TCDD) displays high toxicity in animals and has been implicated in human carcinogenesis. Although TCDD is recognized as potent carcinogens, relatively little is known about their role in the tumor promotion and carcinogenesis. It is known that TCDD can increase of cancer risk from various types of tissue by a mechanism possibly involving the aryl hydrocarbon receptor (AhR) activation. In this study, effects of TCDD on cellular proliferation of normal human skin and lung fibroblasts, Detroit551 and WI38 cells were investigated. In addition, to enhance our understanding of TCDD-mediated carcinogenesis, we have investigated process in which expression of Erk1/2, cyclinD1, oncogene such as Ha-ras and c-myc, and their cognate signaling pathway. TCDD that are potent activators of AhR-mediated activity was found to induce significant increase of cytochrome P4501A1 mRNA expression, suggesting a presence of functional AhR. These results support that CYP1A1 enzyme may be involved in the generation of TCDD-induced toxicity. Moreover mitogen-activated protein kinases (MARKs) phosphorylation and cyclin D1 overexpression are induced by TCDD, which corresponded with the progression of cellular proliferation. However, TCDD did not affected Ha-ras and c-myc mRNA expression. Taken together, it seems that TCDD are could be a part of cellular proliferation in non-tumorigenic normal human cells such as Detroit551 and WI38 cells through the upregulation of MAPKs signaling pathway regulating growth of cell population. Therefore, AhR-activating TCDD could potentially contribute to tumor promotion and Detroit551 and WI38 cells have been used as a detection system of tumorigenic effects of TCDD.

• 한국어류학회지
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• 제29권1호
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• pp.1-12
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• 2017

낙동강 상류 지류인 황지천에서 쉬리(Coreoleuciscus splendidus)와 참쉬리(C. aeruginosa)의 종 간 자연잡종 개체를 채집하였다. 쉬리와 참쉬리의 종 간 잡종 개체는 외부형태 비교와 함께 핵 DNA의 RAG1 유전자(1,334 bp)와 미토콘드리아 DNA인 CO1 유전자(1,551 bp)를 이용한 염기서열 분석을 실시하였다. 외부형태 분석결과 잡종 개체는 등지느러미, 꼬리지느러미 및 뒷지느러미 3곳에서 지느러미 반문의 형태가 쉬리와 참쉬리의 중간 형태를 나타내었다. RAG1과 CO1 유전자를 이용한 분자계통 분석결과 황치천에 분포하는 쉬리속 어류는 쉬리, 참쉬리 두 종과 두 종 간의 잡종 개체군으로 구성되어 있음을 확인하였으며, CO1 유전자의 염기서열 분석결과 순종인 정교배체와 잡종인 상반교배체가 잘 구분되었다. 또한 RAG1 유전자 분석결과 13개의 염기서열 변이를 확인하였고, 잡종 개체는 9개의 염기서열에서 double peaks가 확인되었다. 유전학적 분석과 외부형태 변이 분석에 의해 쉬리와 참쉬리 사이에 잡종화가 발생한 것을 확인하였으나 잡종 F2세대와 잡종 F1 세대의 생식적 격리 여부는 확인하지 못하였다.

• Journal of Microbiology and Biotechnology
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• 제31권8호
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• pp.1154-1162
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• 2021

The transcriptional capacities of target genes are strongly influenced by promoters, whereas few studies have focused on the development of robust, high-performance and cross-species promoters for wide application in different bacteria. In this work, four novel promoters (P_k.rtufB, P_k.r1, P_k.r2, and P_k.r3) were predicted from Ketogulonicigenium robustum and their inconsistency in the -10 and -35 region nucleotide sequences indicated they were different promoters. Their activities were evaluated by using green fluorescent protein (gfp) as a reporter in different species of bacteria, including K. vulgare SPU B805, Pseudomonas putida KT2440, Paracoccus denitrificans PD1222, Bacillus licheniformis and Raoultella ornithinolytica, due to their importance in metabolic engineering. Our results showed that the four promoters had different activities, with P_k.r1 showing the strongest activity in almost all of the experimental bacteria. By comparison with the commonly used promoters of E. coli (tufB, lac, lacUV5), K. vulgare (Psdh, Psndh) and P. putida KT2440 (JE111411), the four promoters showed significant differences due to only 12.62% nucleotide similarities, and relatively higher ability in regulating target gene expression. Further validation experiments confirmed their ability in initiating the target minCD cassette because of the shape changes under the promoter regulation. The overexpression of sorbose dehydrogenase and cytochrome c551 by P_k.r1 and P_k.r2 resulted in a 22.75% enhancement of 2-KGA yield, indicating their potential for practical application in metabolic engineering. This study demonstrates an example of applying bioinformatics to find new biological components for gene operation and provides four novel promoters with broad suitability, which enriches the usable range of promoters to realize accurate regulation in different genetic backgrounds.