• 한국발생생물학회지:발생과생식
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• 제11권3호
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• pp.195-203
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• 2007

자웅이체형 어류의 성결정 및 성분화는 일반적으로 각 개체의 유전형을 따른다. 자연환경에서는 자신이 가진 유전정보의 조절에 따라 성분화 시기에 아로마테이즈 유전자의 발현이 증가하거나 감소하고, 그 결과 스테로이드 호르몬들의 조성이 결정되어 각기 다른 방향으로 성분화하는 것으로 알려져 있다. 본 연구에서는 해산 태생 어류인 조피볼락(Sebasts schlegeli)의 ovarian type aromatase (P450aromA)와 brain type aromatase (P450aromB)의 유전자를 부분적으로 클로닝하여, 이들의 염기서열을 밝혔으며, 각 유전자에 대한 primer를 제작한 후 성분화 기간 중 이 유전자들의 발현을 조사하였다. 조피볼락 아로마테이즈 유전자들은 조사를 시작한 출산 후 35일째에 여러 개체의 머리와 몸에서 각각 발현되었으나, 52일째에는 아로마테이즈 유전자들의 발현 개체수가 현저히 감소하였다. 그리고 출산 59일째에는 발현 개체 수가 다시 증가하였다. P450aromA과 P450aromB의 발현 양상은 전반적으로 유사하였으나, 45일째에는 P450aromB의 발현 개체수가 P450aromA 발현 개체 수보다 훨씬 많았다. 조직학적 분석을 통해 제시된 이 종의 성분화 시기는 지금까지 출산 $50{\sim}65$일 전후로 알려져 왔으나, 본 연구에서 나타난 아로마테이즈 유전자들의 발현 결과를 볼 때, 이 종의 실질적인 성분화는 알려진 것보다 최소한 $1{\sim}2$주 정도 빨리 진행될 가능성이 있다. 본 연구의 결과는 또한 태생어류의 성분화도 난생어류의 성분화에서 보고된 것과 마찬가지로 아로마테이즈의 작용과 밀접하게 연관되어 있다는 사실을 시사한다.

• 한국응용약물학회:학술대회논문집
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• 한국응용약물학회 1995년도 제3회 추계심포지움
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• pp.153-155
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• 1995

To investigate the mechanism of the regulation of cytochrome P450IAl, the 5'-flanking region of a trout cytochrome P4501Al was cloned into the CAT basic expression vector at HindⅢ site. This trout Cytochrome P450IAl upstream DNA containing CAT construct was transfected into Hepa-1 cells .3MC treatment to hepa I cells transfected with trout P450IAl-CAT construct increased CAT protein and mRNA by 2.81 fold when it was compared with that of control. This increase CAT protein and mRNA was decreased by concomitantly treated flavonoids and aminopyrine. The level of CAT protein was 29.2-58.0% of 3MC stimulated CAT protein.

• Journal of Microbiology and Biotechnology
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• 제19권12호
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• pp.1612-1616
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• 2009

Isoflavonoids are a class of phytoestrogens. Isoflavonone synthase (IFS) is responsible for the conversion of naringenin to genistein. IFS is a cytochrome P450 (CYP), and requires cytochrome P450 reductase (CPR) for its activity. Additionally, the majority of cytochrome P450s harbor a membrane binding domain, making them difficult to express in Escherichia coli. In order to resolve these issues, we constructed an inframe fusion of the IFS from red clover (RCIFS) and CPR from rice (RCPR) after removing the membrane binding domain from RCIFS and RCPR. The resultant fusion gene, RCIFS-RCPR, was expressed in E. coli. The conversion of naringenin into genistein was confirmed using this E. coli transformant. Following the optimization of the medium and cell density for biotransformation, $60\;{\mu}M$ of genistein could be generated from $80\;{\mu}M$ of naringenin. This fusion protein approach may be applicable to the expression of other P450s in E. coli.

• 생명과학회지
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• 제34권3호
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• pp.199-207
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• 2024

CYP 효소는 내인성 및 외인성 화학물질의 대사에 핵심적인 역할을 담당한다. 특히, 약물, 자연생성물 및 환경 독성 화학물질 등은 조직에 따라 특이적 CYP 효소 유전자의 발현을 조절하며, 이는 체내 유입된 약물과 독성물질 등과 같은 화학물질들 사이의 상호작용을 유발하여 이들의 대사에 영향을 줌으로써, 결국 치료 효과와 독성 효과에 변화를 초래한다. 이 분야에 대한 지난 수십 년 동안의 집중적인 연구는, 이러한 CYP 효소 유전자의 발현조절을 매개하는 분자적 기전을 이해하는 데 상당한 진전을 가져왔다. 이제는 구체적인 CYP 효소 유전자의 발현 유도를 조절하는 화합물들뿐만 아니라, 이를 감지하여 특정 CYP 유전자의 발현 조절을 매개하는 데 관여하는 수용체들과 이들의 신호전달 경로들이 비교적 상세히 밝혀졌다. 본 총설에서는, 다양한 화학물질의 노출에 반응하여 CYP 효소 유전자들의 발현 유도를 매개하는 것으로 알려진 주요 외인성 물질-감지 수용체들과 기타 조절인자들의 분자적 작용기전을 요약한다.

• Journal of Microbiology and Biotechnology
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• 제13권4호
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• pp.624-627
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• 2003

Benzo[a]pyrene (B[a]P) is an environmental pollutant that has been implicated in carcinogenesis. Saccharomyces cerevisiae was treated with B[a]P, and the responses of its cytochrome P450 (CYP) enzyme and DNA-damage checkpoint genes were examined through gene expression profiles using a reverse transcription polymerase chain reaction (RT-PCR). The DNA-damage checkpoint genes tested were the chk1 and pds1 genes, involved in a metaphase arrest, the swi6 gene targeted by G1 arrest, the pol2 gene related to S phase arrest, and the cln2 gene encoding a cyclin protein, all of which are based on rad9 and rad24. Among these genes, no noticeable effect was found when the cells were exposed to various concentrations of B[a]P. However, the transcriptional activity of CYP51 was significantly different when the cells were exposed to B[a]P. Accordingly, the present results indicate that cytochrome P450 plays a more significant role than DNA-damage checkpoint genes in the response of S. cerevisiae to B[a]P.

• Journal of Microbiology and Biotechnology
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• 제21권1호
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• pp.14-19
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• 2011

A cyclic undecapeptide-family natural product, cyclosporin A (CyA), which is one of the most valuable immunosuppressive drugs, is produced nonribosomally by a multifunctional cyclosporin synthetase enzyme complex in a filamentous fungal strain named Tolypocladium niveum. Previously, structural modifications of cyclosporins such as a regionspecific hydroxylation at the $4^{th}$ N-methyl leucine in a rare actinomycetes called Sebekia benihana were reported to lead to dramatic changes in their bioactive spectra. However, the reason behind this change could not be determined since a system to genetically manipulate S. benihana has not yet been developed. To address this limitation, in this study, we utilized the most commonly practiced gene manipulation techniques including conjugation-based foreign gene transfer-and-expression as well as targeted gene disruption to genetically manipulate S. benihana. Using these optimized genetic manipulation systems, a putative cytochrome P450 hydroxylase (CYP) gene named CYP506, which is involved in CyA hydroxylation in S. benihana, was specifically disrupted and genetically complemented. The S. benihana${\Delta}$CYP506 exhibited a significantly reduced CyA hydroxylation yield as well as considerable yield restoration by functional complementation of the S. benihana CYP506 gene, suggesting that the genetically manipulated S. benihana CYP mutant strains may serve as a more efficient bioconversion host for various valuable metabolites including CyA.

• Journal of Ginseng Research
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• 제43권4호
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• pp.645-653
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• 2019

Background: Cytochrome P450 enzymes catalyze a wide range of reactions in plant metabolism. Besides their physiological functions on primary and secondary metabolites, P450s are also involved in herbicide detoxification via hydroxylation or dealkylation. Ginseng as a perennial plant offers more sustainable solutions to herbicide resistance. Methods: Tissue-specific gene expression and differentially modulated transcripts were monitored by quantitative real-time polymerase chain reaction. As a tool to evaluate the function of PgCYP736A12, the 35S promoter was used to overexpress the gene in Arabidopsis. Protein localization was visualized using confocal microscopy by tagging the fluorescent protein. Tolerance to herbicides was analyzed by growing seeds and seedlings on Murashige and Skoog medium containing chlorotoluron. Results: The expression of PgCYP736A12 was three-fold more in leaves compared with other tissues from two-year-old ginseng plants. Transcript levels were similarly upregulated by treatment with abscisic acid, hydrogen peroxide, and NaCl, the highest being with salicylic acid. Jasmonic acid treatment did not alter the mRNA levels of PgCYP736A12. Transgenic lines displayed slightly reduced plant height and were able to tolerate the herbicide chlorotoluron. Reduced stem elongation might be correlated with increased expression of genes involved in bioconversion of gibberellin to inactive forms. PgCYP736A12 protein localized to the cytoplasm and nucleus. Conclusion: PgCYP736A12 does not respond to the well-known secondary metabolite elicitor jasmonic acid, which suggests that it may not function in ginsenoside biosynthesis. Heterologous overexpression of PgCYP736A12 reveals that this gene is actually involved in herbicide metabolism.

• Fisheries and Aquatic Sciences
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• 제9권2호
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• pp.64-69
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• 2006

Cytochrome P450 (CYP1A) gene expression in the liver and sex steroid levels in plasma were investigated in olive flounder (Paralichthys olivaceus) exposed to tributyltin (TBT) and benzo[a]pyrene (BaP). We constructed a cDNA library and cloned a 230-base sequence encoding partial CYP1A DNA. The CYP1A gene expression level was estimated using northern blotting. Hepatic CYP1A mRNA levels in fish injected with BaP at 10 mg/kg body weight (b.w.) increased for 48 h after injection. However, fish injected with both BaP and TBT at 10 mg/kg b.w. showed no significant changes in CYP1A mRNA level after 48 h. Plasma concentrations of testosterone and $17{\beta}$-estradiol were not significantly different in males and females injected with BaP and TBT. We suggest that TBT-induced suppression of BaP bioactivity should be interpreted with caution in biomonitoring field studies.

• 생명과학회지
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• 제13권6호
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• pp.879-888
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• 2003

추에서 항균 phytoalexin으로 알려진 capsidiol의 생합성을 촉매하는 5-epi-aristolochene hydroxylase는 cytochrome P450 (P450) 억제제인 ancymidol과 ketocornazol에 의해 그 활성이 특이적으로 억제되어, 이 효소가 P450계 효소임을 알 수 있었다. P450 효소가 공통으로 보유하는 염기서열을 지닌 primer를 이용하여 RT-PCR과 cDNA screening을 실시한 결과 고추배양세포에서 elicitor 처리에 의해 강하게 유도되는 cDNA (P450Hy01)를 cloning하였다. 배양세포에 cellulase, arachidonic acid, jasmonic acid, 자외선 등을 처리하여 capsidiol을 생합성하는 량과 P450Hy01 mRNA의 발현정도는 밀접한 유사성이 있었다. P450Hy01의 염기서열은 담배에서 밝혀진 5-epi-aristolochene-1,3-hydroxylase와 98%의 유사성이 있었으며, P450 효소가 공통으로 지니는 heme-binding domain인 FxxGxRxCxG를 보유하고 있었다 이러한 결과는 본 연구에서 cloning된 P450Hy01이 고추의 세포에서 capsidiol의 생합성을 촉매하는 5-epi-aristolochene hydroxylase 효소를 coding 하고 있음을 시사한다.

• 대한약학회:학술대회논문집
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• 대한약학회 2003년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2-2
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• pp.127.2-128
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• 2003

Sepsis remains common surgical problems with high morbidity and mortality despite improvement in the management for septic patient. Although hepatocellular dysfunction occurs during sepsis, the mechanism responsible for this remains unclear. In sepsis, a state of severe oxidative stress is encountered, with host endogenous antioxidant defenses overcome. Therefore, the aim of this study was to determine whether specific abnormality exists in cytochrome P450 (CYP)-mediated metabolizing function associated with polymicrobial sepsis and whether role of ascorbic acid (AA) in the alterations during sepsis. (omitted)