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- 저자 : Sasame, Henry A. (검색결과 1 건)
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Ritter, Joseph K.,Owens, Ida S.,Negishi, Masahiko,Nagata, Kiyoshi,Sheen, Yhun Y.,Gillette, James R.,Sasame, Henry A. 이화여자대학교 생명과학연구소 1991 생명과학연구논문집 Vol.2 No.-
We have isolated a cDNA clone, Nah-2, encoding the cytochrome P-450^Nah(naphthalene hydroxylase) from a mouse lung λZAP cDNA library using anti-cytochrome P-450^Nah IgG as a probe. This same antibody selectively blocked[Nagata, K.,Martin, B. M., Gillette, J. R., & Sasame, H. A.(1990) Drug Metab. Dispos. 18, 557-564] the cytochrome P-450 in mouse lung microsomes that catalyzed the conversion of naphthalene to (1R,2S)- naphthalene 1,2-oxide, which has been postulated as a causative agent in the naphthalene-induced tissue-specific necrosis of Clara cells in mouse lung. The toxic effect is seen in mouse and not in rat. The cDNA encodes a polypeptide of 491 amino acids with a molecular mass of 50 kDa. Northern blot analysis with an Nah-2specific probe revealed that the mRNA is expressed in a species-and tissue-specific manner, present only in mouse lung and liver and not in that of rat. The mRNA encoding Nah-2 is constitutively expressed and is not induced by either phenobarbital, pyrazole, pregnenolone 16∂-carbonitrile, or 3-methylcholanthrene. Comparative amino acid sequence analyses with other documented members of the P-450 gene superfamily revealed that this encoded protein is in the IIF subfamily. To analyze its substrate specificity, the cDNA was inserted into the vector, pAAH5, and expressed in the Saccharonyces cerevisiae strain, AH_22. The presence of cytochrome P-450_Nah in the micreosomes isolated from transformed cell and analyzed by Western blot was confirmed by immunocomplexing product with anti-cytochrome P450_Nah IgG. Furthermore, activity toward naphthalene in the microsomes from the transformed cells established that this clone encodes a naphthalene hydroxylase. Like lung microsomes and purified and reconstituted cytochrome P450_Nah, transformed yeast microsomes convert naphthalene primarily to the trans-(1R)-hydroxy-(2R)-glutathionyl-1, 2-dihydronaphthalene conjugate, a stable form of the putative toxicant(1R,2S) oxide in the presence of glutathione and a mixture of glutathione S-transferases. Results of immunochemical studies support a role of this cytochrome P-450 in lung toxicity in mice exposed to high doses of naphthalene.
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