• Korean Journal of Environmental Biology
• /
• v.24 no.2 s.62
• /
• pp.195-200
• /
• 2006

The response of hepatic mixed function oxygenase (MFO) system was investigated in olive flounder exposed to formalin. Hepatic microsome of olive flounder incubated in vitro with formalin demonstrated the induction of cytochrome P450 (CYP), ethoxyresorufin deethylase (EROD), cytochrome P450 reductase (P450R) and cytochrome b5 reductase (b5R) activity. In addition, olive flounder was exposed to 100, 300 and 500 ppm of formalin for 1 h and then transferred to a flow-through type of 1000 L aquarium. Hepatic MFO enzyme activity was determined for 72 h. As the result, hepatic CYP, P450R and EROD activities increased following exposure of formalin, but b5R and GST showed no significant change. These results imply that CYP and P450R can be considered as main hepatic enzymes involving in detoxification of formalin.

• Journal of Embryo Transfer
• /
• v.12 no.1
• /
• pp.49-56
• /
• 1997

This study was carried out to establish an effective and practical system for commercialization of embryo production techniques by analyzing several factors influencing in vivo embryo production on condition of donors treated FSH-P and SUPER-OV in Korean native cattle. In vivo embryos were flushed 226 times from 128 donors. The results obtained for the factors influencing in vivo embryo production by conditionof donors treated FSH-P and SUPER-OV were as follows :1. Age and parity of donors did not affect the transferable and freezable embryos among the treatments in FSH-P but the transferable and freezable embryos were decreased after 6 years old and 4th parity in SUPER-OV(P<0.01). 2. The production of embryos on the weight of donors was higher in over 400kg of body weight in FSH-P(P<0.01) and was higher in over 450kg than 400~450kg of body weight in SUPER-OV(P<0.05). For FSH-P embryo production was better responded in 350~450kg of body weight with 30~32mg doses, and showed a better result in over 450kg body weight with 32~34mg doses.(Key Words : in vivo embryo, donors, FSH-P SUPER-OV)

• Microbiology and Biotechnology Letters
• /
• v.47 no.4
• /
• pp.536-545
• /
• 2019

Cytochrome P450 (P450 or CYP) is involved in the metabolism of endogenous and exogenous compounds in most organisms. P450s have great potential as biocatalysts in the pharmaceutical and fine chemical industries because they catalyze diverse oxidative reactions using a wide range of substrates. The high-cost nicotinamide cofactor, NADPH, is essential for P450 reactions. Glucose-6-phosphate dehydrogenase (G6PDH) has been commonly used in NADPH-generating systems (NGSs) to provide NADPH for P450 reactions. Currently, only two G6PDHs from Leuconostoc mesenteroides and Saccharomyces cerevisiae can be obtained commercially. To supply high-cost G6PDH cost-effectively, we cloned the cytosolic G6PDH gene of Solanum lycopersicum (tomato) with 6xHis tag, expressed it in Escherichia coli, and purified the recombinant G6PDH (His-G6PDH) using affinity chromatography. In addition, enzymatic properties of His-G6PDH were investigated, and the His-G6PDH-coupled NGS was optimized for P450 reactions. His-G6PDH supported CYP102A1-catalyzed hydroxylation of omeprazole and testosterone by NADPH generation. This result suggests that tomato His-G6PDH could be a cost-effective enzyme source for NGSs for P450-catalyzed reactions as well as other NADPH-requiring reactions.

• Asian Pacific Journal of Cancer Prevention
• /
• v.15 no.16
• /
• pp.6575-6580
• /
• 2014

This study aimed to evaluate the antimutagenic and anticarcinogenic activity of turmeric essential oil as well as to establish biochemical mechanisms of action. Antimutagenicity testing was accomplished using strains and known mutagens with and without microsomal activation. Anticarcinogenic activity was assessed by topical application of 7, 12 - dimethylbenz[a]anthracene (DMBA) as initiator and 1% croton oil as promoter for the induction of skin papillomas in mice. Inhibition of p450 enzymes by TEO was studied using various resorufins and aminopyrene as substrate. Turmeric essential oil (TEO) showed significant antimutagenic activity (p<0.001) against direct acting mutagens such as sodium azide ($NaN_3$), 4-nitro-O-phenylenediamine (NPD) and N-methyl-N-nitro N'nitrosoguanine (MNNG). TEO was found to have significant antimutagenic effect (>90%) against mutagen needing metabolic activation such as 2-acetamidoflourene (2-AAF). The study also revealed that TEO significantly inhibited (p<0.001) the mutagenicity induced by tobacco extract to Salmonella TA 102 strain. DMBA and croton oil induced papilloma development in mice was found to be delayed and prevented significantly by TEO application. Moreover TEO significantly (P<0.001) inhibited isoforms of cytochrome p450 (CYP1A1, CYP1A2, CYP2B1/2, CYP2A, CYP2B and CYP3A) enzymes in vitro, which are involved in the activation of carcinogens. Results indicated that TEO is antimutagenic and anticarcinogenic and inhibition of enzymes (p450) involved in the activation of carcinogen is one of its mechanisms of action.

• Archives of Pharmacal Research
• /
• v.27 no.2
• /
• pp.232-238
• /
• 2004

Trolox is a hydrophilic analogue of vitamin E. The aim of this study was to investigate its effects on hepatic injury, especially alteration in cytochrome P450 (CYP)-dependent drug metabolism during polymicrobial sepsis. Rats were subjected to polymicrobial sepsis by cecal ligation and puncture (CLP). The rats were treated intravenously with Trolox (2.5 mg/kg) or vehicle, immediately after CLP. Serum aminotransferases and lipid peroxidation levels were markedly increased 24 h after CLP. This increase was attenuated by Trolox. Total CYP content and NADPH-P450 reductase activity decreased significantly 24 h after CLP. This decrease in CYP content was attenuated by Trolox. At 24 h after CLP, there was a significant decrease in the activity of these CYP isozymes: CYP1A1, 1A2, 2B1, and 2E1. However, Trolox differentially inhibited the decrease in CYP isozyme activity. Trolox had little effect on the decrease in CYP1A1 activity but Trolox significantly attenuated decreases in CYP1A2 and 2E1 activities. In fact, Trolox restored CYP2B1 activity to the level of activity found in control rats. Our findings suggest that Trolox reduces hepatocellular damage as indicated by abnormalities in hepatic drug-metabolizing function during sepsis. Our data also indicates that this protection is, in part, caused by decreased lipid peroxidation.

• Journal of Microbiology and Biotechnology
• /
• v.27 no.5
• /
• pp.983-989
• /
• 2017

NADPH-P450 reductase (NPR) transfers electrons from NADPH to cytochrome P450 and heme oxygenase enzymes to support their catalytic activities. This protein is localized within the endoplasmic reticulum membrane and utilizes FMN, FAD, and NADPH as cofactors. Although NPR is essential toward enabling the biochemical and pharmacological analyses of P450 enzymes, its production as a recombinant purified protein requires a series of tedious efforts and a high cost due to the use of $NADP^+$ in the affinity chromatography process. In the present study, the rat NPR clone containing a $6{\times}$ Histidine-tag (NPR-His) was constructed and heterologously expressed. The NPR-His protein was purified using $Ni^{2+}$-affinity chromatography, and its functional features were characterized. A single band at 78 kDa was observed from SDS-PAGE and the purified protein displayed a maximum absorbance at 455 nm, indicating the presence of an oxidized flavin cofactor. Cytochrome c and nitroblue tetrazolium were reduced by purified NPR-His in an NADPH-dependent manner. The purified NPR-His successfully supported the catalytic activities of human P450 1A2 and 2A6 and fungal CYP52A21, yielding results similar to those obtained using conventional purified rat reductase. This study will facilitate the use of recombinant NPR-His protein in the various fields of P450 research.

• Toxicological Research
• /
• v.27 no.1
• /
• pp.25-29
• /
• 2011

The cytochrome P450 (P450, CYP) are the superfamily of heme-containing monooxygenase enzymes, found throughout all nature including mammals, plants, and microorganisms. Mammalian P450 enzymes are involved in oxidative metabolism of a wide range of endo- and exogenous chemicals. Especially P450s involved in drug metabolisms are important for drug efficacy and polymorphisms of P450s in individuals reflect differences of drug responses between people. To study the functional differences of CYP2A6, CYP2D6, and CYP4A11 variants, we cloned the four CYP2A6, three CYP2D6, and three CYP4A11 variants, which were found in Korean populations, in mammalian expression vector pcDNA by PCR and examined their expressions in COS-7 mammalian cells using immunoblots using P450 specific polyclonal antibodies. Three of four CYP2A6, two of three CYP4A11, and two of three CYP2D6 variants showed expressions in COS-7 cells but the relative levels of expressions are remarkably different in those of each variants. Our findings may help to study and explain the differences between functions of CYP variants and drug responses in Korean populations.

• Journal of Microbiology and Biotechnology
• /
• v.20 no.4
• /
• pp.712-717
• /
• 2010

Cytochrome P450 enzymes (P450s) are involved in the synthesis of a wide variety of valuable products and in the degradation of numerous toxic compounds. The P450 BM3 (CYP102A1) from Bacillus megaterium was the first P450 discovered to be fused to its redox partner, a mammalian-like diflavin reductase. Here, we report the development of a whole-cell biocatalyst using ice-nucleation protein (Inp) from Pseudomonas syringae to display a hemeand diflavin-containing oxidoreductase, P450 BM3 (a single, 119-kDa polypeptide with domains of both an oxygenase and a reductase) on the surface of Escherichia coli. The surface localization and functionality of the fusion protein containing P450 BM3 were verified by flow cytometry and measurement of enzymatic activities. The results of this study comprise the first report of microbial cell-surface display of a heme- and diflavin-containing enzyme. This system should allow us to select and develop oxidoreductases containing heme and/or flavins into practically useful whole-cell biocatalysts for extensive biotechnological applications, including selective synthesis of new chemicals and pharmaceuticals, bioconversion, bioremediation, live vaccine development, and biochip development.

• Journal of Sasang Constitutional Medicine
• /
• v.24 no.4
• /
• pp.84-91
• /
• 2012

Objectives : The purpose of this study is to investigate the inhibitory or inductive potentials of Yuldahanso-tang (YDT) and Chungsimyonja-tang (CST), herbal formulas for Taeeumin, on cytochrome P450 (CYP450) drug metabolizing enzyme. The mechanisms for the herbal formula-drug interaction has not been well reported in spite of the chance for co-administration with conventional drugs. Methods : To evaluate the interaction potential of YDT-drug or CST-drug, the fluorescence-based enzyme assays on CYP450 isozymes including CYP3A4, CYP2C19, CYP2D6 and CYP2E1 were established in vitro. The inhibitory effects of herbal formulas were characterized with $IC_{50}$ values. Results : YDT showed inhibitory effects on CYP2D6 and CYP2E1-mediated metabolism, while it exhibited week inhibition on CYP3A4 and CYP2C19 relatively. CST exerted relatively weak inhibitory effects on the four CYP450 isozymes compared to that of YDT. Conclusions : These results suggest that the herbal formula-drug interaction could be occur when YDT are co-administered with drugs mediated by CYP2D6 or CYP2E1.

• Journal of Microbiology and Biotechnology
• /
• v.11 no.6
• /
• pp.1011-1017
• /
• 2001

Streptomyces sp. Y-110 cytochrome P-450, induced by the addition of compactin -Na into the culture medium, was purified from the cell extract to apparent homogeniety, mainly by DEAE-Sepharose, hydroxyapatite, and Mono Q column chromatyography. The sepcific activity of purified enzyme on its substrate, compactin-Na, was determined to be 15 nmol of pravastatin per mg protein. The molecular mass of this enzyme on SDS-PAGE was $37{\pm}0.5$ kDa, pI was 4.5, and its CO difference spectrum showed maximum absorption peaks at 452 and 550nm, respectively. The N-terminal amino acid sequence was determined to be Met>Thr>Cys>Thr>Pro>Val>Thr>Val>The>Gly>Ala>Ala>Gly>Gln>Ile>Gly>Tyr>Ala>Leu. Its apparent $K_m$ on compactin-Na was $1.294{\mu}M{\cdot}min^-1,\;and\;V_{max}\;was\;1.028{\mu}M{\cdot}min^-1$. The maximum substrate concentration ($K_s$) for reaction was $270 {\mu}M$and thus $1/[K_s]$ was $3.7{\mu}M$. These physicochemical characteristics and kinetic behavior of this enzyme were compared and shown to be different from those of Streptomyces cytochrome P-450 enzymes reported, suggesting that this enzyme may be an additional member of the Streptomyces cytochrome P-450 family.