• Journal of Microbiology and Biotechnology
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• v.15 no.1
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• pp.188-191
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• 2005

Abstract Cyclosporins are a family of clinically-important immunosuppressive cyclic peptides produced by Tolypocladium inflatum. The structural modification of cyclosporins via hydroxylation at various positions of N-methyl leucines in cyclosporin A leads to a dramatic change of their bioactive spectra. Among over 100 soil actinomycetes screened, two actinomycetes species, Sebekia benihana and Pseudonocardia autotrophica, were identified to contain superior cyclosporin A hydroxylation activities. A HPLC-based cyclosporin A hydroxylation assay revealed that each strain possesses distinctive hydroxylation specificity and regiospecificity; mono-hydroxylation at the 4th N-methyl leucine of cyclosporin A by S. benihana, and di-hydroxylations at both 4th and 9th N-methyl leucines of cyclosporin A by P. autotrophica. The conversion yields for cyclosporin A hydroxylation by both S. benihana and P. autotrophica were significantly improved from less than 10% and 18% up to 58% and 45%, respectively, in the optimized culture containing molybdenum with 0.05 g/l of cyclosporin A concentration. An ancymidol-specific inhibition of cyclosporin hydroxylation also suggested that the regiospecific cyclosporin hydroxylation might be catalyzed by a putative cytochrome P450 mono-oxygenase enzyme.

• BMB Reports
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• v.32 no.3
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• pp.226-231
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• 1999

In vitro effects of acetone on cytochrome P450 (P450)-dependent benzo(a)pyrene (B(a)P) hydroxylation supported by cumene hydroperoxide (CuOOH) or NADPH/$O_2 $ systems were studied using 3-methylcholanthrene-pretreated rat liver microsomes. The maximal rate of B(a)P hydroxylation at constant concentration ($80\;{\mu}M)$ of the substrate was observed in the presence of $30\;{\mu}M$ CuOOH. However, at concentrations higher than $30\;{\mu}M$ CuOOH the hydroxylation rates were rapidly decreased. In contrast to CuOOH, at a concentration of $200\;{\mu}M$ NADPH, B(a)P hydroxylation rate reached a plateau. At concentrations higher than $200\;{\mu}M$ NADPH, the rates of substrate hydroxylation were maintained at the maximal rate with no inhibition. Acetone at 1% (v/v) enhanced both CuOOH- and NADPH/$O_2$-supported B(a)P hydroxylation at the optimal concentrations of the cofactors. At concentrations higher than 1% (v/v) acetone, substrate hydroxylation was sterero specific under the support of these two cofactors; it was strongly enhanced with $30\;{\mu}M$ CuOOH, but rather inhibited in the $200\;{\mu}M$> NADPH/$0_2 $ system. The lipid peroxidation rate induced during CuOOH-supported P450-dependent B(a)P hydroxylation was increased as CuOOH concentrations were increased. Acetone in the concentration range of 2.5~7.5%(v/v) inhibited lipid peroxidation during CuOOH supported B(a)P hydroxylation. The finding that CuOOH-supported B(a)P hydroxylation is greatly enhanced by acetone suggests that acetone may contribute more to the activation of oxygen (for the insertion of oxygen into the substrate) in the presence of CuOOH than with NADPH/$O_2$. Acetone may also contribute to the partial inhibition of destruction of microsomal membranes by lipid peroxidation.

• Archives of Pharmacal Research
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• v.15 no.1
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• pp.41-47
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• 1992

In order to obtain acetaminophen, a popular analgesic-antipyretic, though microbial p-hydroxylation and N-acetylation of aniline, various Streptomyces strains were screened. Aniline N-acetylation activity was rather ubiquitous but-hydroxylation activity was selective. Microbial conversion pathway of aniline to acetaminophen was considered to be through N-acetylation and p-hydroxylation or vice versa. However, depending on species used, o-hydroxylation and its degradation activity (S. fradiae) and acetaminophen degradation activity (S. coelicolar) were also detected. Among the screened Streptomyces strains, S fradiae NRRL 2702 showed the highest acetanilide p-hydroxylation activity (203% conversion rate). Furthermore, in S. fradiae carbon source and its concentration, phosphate ion concentration and pH of growth medium were found to play the crucial roles in p-hydroxylation activity. Through the proper combination of factors mentioned above, the ten times more activity (26-30% conversion rate) was attained.

• Archives of Pharmacal Research
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• v.15 no.1
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• pp.30-34
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• 1992

In order to obtain acetaminophen, a popular analgesic-antipyretic, through microbial p-hydroxylation and N-acetylation of aniline, various fungi and bacteria were secreened. Among them, Streptomyces species were chosen for strain improvement by the use of interspecific protoplast fusion technique. Two interspecific fused strains were developed between S. rimosus (N-cetylation function) and S. aureofaciens (p-hydroxylation function) and also between S. lividans and S. globisporus. For efficient protoplast fusion and cell wall regeneration, various conditions were examined. In a typical experiment of mixed S rimosus ($pro^- \;his^-$) and S. aureofaciens ($ilv^-$) protoplasts with 40% (w/v) polythylene glycol 3350 (PEG) for 3 min gave $8.3\times10^{-7}$ of fusion frequency. Treatment of mixed S. lividans (pant-) and S. globisporus (leu-) protoplasts with 50% (w/v) PEG for 3 min at $30^\circ{C}$ gave $1.2\times10^{-6}$ of frequency. Among the fused strains, up to 40-50% increase in p-hydroxylation power was observed. To investigate the possibility of plasmid involvement in p-hydroxylation power was observed. To investigate the possibility of plasmid involvement in p-hydroxylation of acetanilide, plasmid curing was attempted. We found that cells treated with acriflavine (at the frequency of 100%) and cells regenerated from protoplsts of S. auroefaciens (2% frequency) lost their p-hydroxylation function.

• Korean Journal of Microbiology
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• v.27 no.4
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• pp.366-372
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• 1989

Twenty-one strains were tested for 11$\beta$-hydroxylation of Reichstein's substance S. Four fungi exhibited ability for the reaction, among which Pellicularia fillamentosa showed the highest activity. The 11$\beta$-hydroxylase of this fungus was proved to be induced by the substrate, cycloheximide reducing significantly the activity of the enzyme. Range of optimum pH for the 11$\beta$-hydroxylation was broad and found to be 2.0-8.0. Test of the enzyme activity at different growing stages, from spore to mycelia, showed that the branching stage of hyphae and the mature mycelial stage were the most effective for the Reichstein's substance S transformation. However, 11$\beta$-hydroxylase in the intact spore was turned out to be uninducible with the substrate.

• Korean Journal of Microbiology
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• v.31 no.4
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• pp.306-311
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• 1993

The effects of methanol. used as a solvent for the hydrophobic substrate progesterone. on the morphology of Rhizopus nigricans and 11$\alpha$-hydroxylation of progesterone was investigated. The methanol tolerance of the 11$\alpha$-hydroxylase system in polyacrylamide immobilized R. nigricans mycelia as well as in free mycelia has been induced by adding various unsaturated fatty acids. biotin and ions into the cultivation medium. Immobilization of the cell seemed to protect the cells from denaturation by methanol. It gave higher reaction rate of progesterone than the free mycelia in the presence of methanol.500 $\mu$g/l of biotin was found to be the most effective induction agent for the methanol tolerance among tested chemicals. R. nixricans cells sustained its enzymatic activity at higher methanol concentrations as a result of accumulation of unsaturated fatty acids. especially oleic acid. in the membrane phospholipid.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1994.04a
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• pp.334-334
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• 1994

Omeprazole의 약동학적 Parameter는 tmax를 제외하고 두 군간에 유의한 (p<0.001) 차이론 보였다 : AUC의 평균치는 5-mephenytoin hydroxylation poor metabol izer에서 extensive metabol izer에 비해 약 6-7배 컸다. Omeprazole sulfone의 parameter는 omeprazole에서 관찰된 두 군간의 차이와 유사하였다. 그러나 5-hydroxyomeprazole의 경우에는 Cmax, AUC 등이 extensive metabolizer에서 더컸다. Omeprazole의 청소율은 S-mephenytoin hydroxylation 대사능(MR)과 유의한 상잔관계(rs=0.79, p<0.01)를 보였으며, omeprazole과 그 대사물(5-hydroxyomeprazole 및 omeprazole sulfone)의 반감기 또한 S-mephenytoin hydroxylation 대사능과 유의한 상관성을 보였다.

• Archives of Pharmacal Research
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• v.15 no.1
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• pp.35-40
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• 1992

S. fradiae showed the highest acetanilide p-hydroxylation activity in the tested strains. And S. fradiae was well characterized genetically, especially with respect to tylosin production. Two mutants, which lost hydroxylation, were isolated in 140 regenerated colonies from protoplasts. In restriction enzyme digesion of total DNAs, isolation of giant linear plasmid DNA and determination of antibiotic resistances to chloramphenicol, tylosin, hygromycin B and mitomycin C, any differences among mutants and a wild type strain were not detected. These facts suggest that lesion on 6, 000 Kb chromosomal DNA was responsible for the lack of p-hydroxylation activity induced by protoplast formation and regeneration.

• Bulletin of the Korean Chemical Society
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• v.22 no.1
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• pp.93-96
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• 2001

• Korean Journal of Microbiology
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• v.29 no.4
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• pp.258-262
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• 1991

Rate of 11.alpha.-hydroxylation of progesterone with Rhizopus nigricans was accelerated by glucose. Glucose seemed to play an important role in the formation of cofactor because its effects on the reaction were almost same as those of electron acceptors such as NADPH and $NaIO_{4}$. Rate of glucose consumption appeared to increase in proportion as the rate of hydroxylation reaction, which enhanced with increase in the glucose concentration to level off at 0.5 g/l for mycelia and at 20 g/l for spores. However, for mycelia immobilized in polyacrylamide gel, externally added glucose did not affect the reaction rate at all because of the glucose accumulated in the gel during the cultivation period. 5.alpha.-Reduction of 11.alpha.-hydroxyprogesternoe required much higher concentration of glucose than 11.alpha.-hydroxylation of progesternoe so that high yield of 11.alpha.-hydroxyprogesternoe can be obtained by repressing the activity of 5.alpha.-reductase at low concentration of glucose.