|Budget Amount *help
¥2,900,000 (Direct Cost : ¥2,900,000)
Fiscal Year 2000 : ¥600,000 (Direct Cost : ¥600,000)
Fiscal Year 1999 : ¥1,200,000 (Direct Cost : ¥1,200,000)
Fiscal Year 1998 : ¥1,100,000 (Direct Cost : ¥1,100,000)
Cytochrome P450 (P450) are widely distributed in the biosphere and catalyze a remarkably broad range of chemical reactions on an equally broad range of substrates. Therefore, the P450-producing bacteria are good candidates for biodegradation of hazardous organochemicals.
In this study, 10 strains of P450-producing bacteria were isolated from M9 medium containing a P450-inducer as the sole carbon source from the environment. Strains EP1 to EP7 utilizing 2-ethoxyphenol as the sole carbon source were isolated from the soil of a weed-filled field, the water of a paddy field, laboratory effluent, domestic effluent, river water and activated sludge. Strain MP1 and MP2 utilizing 2-methoxyphenol were isolated from oil-polluted soil and activated sludge, respectively. Strain CP1 utilizing camphor was isolated from the soil of a weed-filled field. This suggests the distribution of P450-producing bacteria in the environment. Metyrapone (2-methyl-1, 2-di-3-pyridyl-1-propanone) inhibited the growt
h of P450-producing bacteria on the media containing a P450-inducer as the sole carbon source, strongly suggesting that the P450 is involved in the catabolism of the inducer.
2-Ethoxyphenol-induced P450EP1A from Rhodococcus sp. strain EP1 and P450EP4 from Gordonia sp. strain EP4 were purified to apparent homogeneity. The P45OEP1A and P450EP4 showed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with molecular weight of about 45 kDa and 47kDa, respectively. The purified P450 degraded 2-ethoxyphenol in an assay system composed of spinach ferredoxin, spinach ferredoxin oxidoreductase and NADPH.In addition to 2-ethoxyphenol, the P450EP1A metabolized 2-methoxyphenol, 1, 1, 1-trichloroethane, carbon tetrachloride, benzene and toluene. P450EP4 metabolized not only 2-ethoxyphenol but also trichloroethylene, dichloroethane, 1, 1, 1-trichloroethane and chloroform. These P450s can display a remarkable chemical flexibility toward substrate. Thus, P450EP1A and P450EP4 may be useful for the removal of hazardous organochemicals from environment.
P450精製品の分光学的性質の検討を行った結果,P450に特徴的な性質を示した。また,P450精製品とほうれん草のフェレドキシン及びフェレドキシンレダクターゼを用いて代謝再構成系を確立し,種々の有害有機化合物の代謝活性を測定した。P450EP1Aは1,1,1-trichloroethane,carbon tetrachloride,benzene,tolueneを代謝した。P450EP4はtrichloroethylene,dichloroethane,1,1,1-trichloroethane及びchloroformを代謝した。両P450は有機塩素系化合物などの有害物質を代謝できる有用なP450である可能性が示唆された。 Less