Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes.
Purification and characterization of two unique forms of cytochrome P-450 from rabbit nasal microsomes.
复制标题
从兔鼻微粒体中纯化和表征两种独特形式的细胞色素 P-450。
DOI：
10.1021/bi00422a007
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发表时间：
1988
期刊：
影响因子：
2.9
通讯作者：
Coon,MJ
中科院分区：
文献类型：
--
作者：
Ding,XX;Coon,MJ
密歇根大学医学院生物化学系，安娜堡，密歇根州 48109 收稿日期：1988 年 4 月 14 日；修订稿于 1988 年 6 月 28 日收到摘要：两种形式的细胞色素 P-450，命名为 P-450NMa 和 P-450NMb，从兔鼻微粒体中纯化至电泳均质。纯化的细胞色素含有14-16 nmol P-450/mg蛋白质，表观单体分子量分别为49 500和51000。根据多项标准（包括氨基酸组成、吸收光谱和肽图）表明，P-450 的两种鼻腔形式彼此不同。此外，根据 NH2 末端氨基酸序列判断，它们与迄今为止描述的所有其他 P-450 细胞色素不同。在三价铁形式中，P-450NMa 处于低自旋状态，而 P-450NMb 主要处于高自旋状态。当用 NADPH-细胞色素 P-450 还原酶和磷脂重构时，P-450NMa 在乙醇和几种鼻致癌物的氧化中非常活跃，包括 7V-亚硝基二乙胺的 N-脱乙基化、非那西丁的 O-脱乙基化和六甲基磷酰胺的 N-脱甲基化。 P-450NMb 也代谢这些底物，但速率较低。两种鼻腔形式也对睾酮有活性，P-450NMa 氧化 17 位底物产生雄烯二酮，P-450NMb 催化 15a-、16a-和 19-位羟基化。这两种细胞色素占鼻微粒体中总 P-450 的主要部分，但在肝微粒体中无法检测到相应的形式。细胞色素 P-450 参与多种外源物质和内源性物质的氧化代谢是众所周知的。根据最近的综述（Black & Coon，1986、1987），已有 60 多种 P-450 从不同物种（大部分来自肝微粒体）中纯化至电泳均质。其中一些同工酶的组织特异性分布已经确定。例如，P-450 同工酶在免疫化学上与
Department of Biological Chemistry, Medical School, The University of Michigan, Ann Arbor, Michigan 48109 Received April 14, 1988; Revised Manuscript Received June 28, 1988 abstract: Two forms of cytochrome P-450, designated P-450NMa and P-450NMb, were purified to electrophoretic homogeneity from rabbit nasal microsomes. The purified cytochromes, which contained 14-16 nmol of P-450/mg of protein, exhibited apparent monomeric molecular weights of 49 500 and 51000, respectively. As indicated by several criteria, including the amino acid composition, absorption spectra, and peptide maps, the two nasal forms of P-450 are distinct from each other. Furthermore, as judged by the NH2-terminal amino acid sequences, they are distinct from all other P-450 cytochromes described to date. In the ferric form, P-450NMa is in the low-spin state, whereas P-450NMb is predominantly in the high-spin state. When reconstituted with NADPH-cytochrome P-450 reductase and phospholipid, P-450NMa is very active in the oxidation of ethanol as well as several nasal procarcinogens, including the N-deethylation of 7V-nitrosodiethylamine, the O-deethylation of phenacetin, and the N-demethylation of hexamethyl-phosphoramide. P-450NMb also metabolizes these substrates, but at lower rates. Both nasal forms are also active with testosterone, with P-450NMa oxidizing the substrate in the 17-position to give androstenedione and P-450NMb catalyzing hydroxylation in the 15a-, 16a-, and 19-positions. The two cytochromes represent the major portion of the total P-450 in nasal microsomes, but the corresponding forms could not be detected in hepatic microsomes.The involvement of cytochrome P-450 in the oxidative me-tabolism of numerous xenobiotics as well as endogenous substances is well recognized. As reviewed recently (Black & Coon, 1986, 1987), more than 60 P-450s have been purified to electrophoretic homogeneity from various species, mostly from liver microsomes. The tissue-specific distribution of some of these isozymes has been established. For example, P-450 isozymes that are immunochemically indistinguishable from