PURIFICATION AND CHARACTERIZATION OF LIVER MICROSOMAL CYTOCHROMES-P-450 - ELECTROPHORETIC, SPECTRAL, CATALYTIC, AND IMMUNOCHEMICAL PROPERTIES AND INDUCIBILITY OF 8 ISOZYMES ISOLATED FROM RATS TREATED WITH PHENOBARBITAL OR BETA-NAPHTHOFLAVONE
PURIFICATION AND CHARACTERIZATION OF LIVER MICROSOMAL CYTOCHROMES-P-450 - ELECTROPHORETIC, SPECTRAL, CATALYTIC, AND IMMUNOCHEMICAL PROPERTIES AND INDUCIBILITY OF 8 ISOZYMES ISOLATED FROM RATS TREATED WITH PHENOBARBITAL OR BETA-NAPHTHOFLAVONE
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DOI：
10.1021/bi00266a045
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发表时间：
1982-01-01
期刊：
影响因子：
2.9
通讯作者：
KAMINSKY, LS
中科院分区：
文献类型：
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作者：
GUENGERICH, FP;DANNAN, GA;KAMINSKY, LS
用一种常见的方法，从大鼠经苯巴比妥或-萘黄酮处理的肝微粒体中纯化出8种不同形式的细胞色素P-450 (P-450)，以达到电泳均匀性。在家兔中制备了针对其中7种病毒的抗体。8 P-450通过铁、铁和铁羰基形式的光谱性质、表观单体MW肽定位、免疫反应性（通过双扩散免疫沉淀分析和交叉免疫电泳识别）、对底物乙酰苯胺、氨基吡啶、苯胺、苯并[a]芘、d-苯并苯胺、N，N''-二甲基亚硝胺、7-乙氧基香豆素、7-乙氧基间苯甲酚、乙基吗啡、对硝基苯甲醚、睾酮和(R)-和(S)-华法林采用交叉十二烷基硫酸钠-聚丙烯酰胺凝胶免疫电泳法对未处理大鼠和经苯巴比妥、5,6-苯甲黄酮、孕烯醇酮-16 - α处理的大鼠肝微粒体中8种形式P-450的含量进行了估计。-碳腈、异黄樟醇或多氯联苯混合物Aroclor 1254。在每种情况下，这8种P-450含量的总和至少与光谱测定的P-450含量一样高。不同的化合物可以诱导不同形式的P-450，单一化合物可以降低一种形式的P-450的水平，同时诱导一种或多种其他形式的P-450。用微粒体制剂观察到的对每种底物的催化活性与基于每种8 P-450含量预测的速率进行了比较。这些研究为进一步研究个体P-450的调控、不同P-450的物理性质以及P-450形态变化在大鼠肝脏模型中的代谢后果提供了基础。
Eight different forms of cytochrome P-450 (P-450) were purified to electrophoretic homogeneity, by a common procedure, from liver microsomes of rats treated with phenobarbital or .beta.-naphthoflavone. Antibodies were prepared to 7 of these forms in rabbits. The 8 P-450 were distinguished by spectral properties of the ferric, ferrous and ferrous carbonyl forms, apparent monomeric MW peptide mapping, immunological reactivity, as discerned by double-diffusion immunoprecipitin analysis and crossed immunoelectrophoresis, and catalytic activities toward the substrates acetanilide, aminopyrine, aniline, benzo[a]pyrene, d-benzphentamine, N,N''-dimethylnitrosamine, 7-ethoxycoumarin, 7-ethoxyresorufin, ethylmorphine, p-nitroanisole, testosterone and (R)- and (S)-warfarin. Crossed sodium dodecyl sulfate-polyacrylamide gel immunoelectrophoresis was used to estimate the levels of each of the 8 forms of P-450 present in the liver microsomes of untreated rats and rats treated with phenobarbital, 5,6-benzoflavone, pregnenolone-16.alpha.-carbonitrile, isosafrole or the polychlorinated biphenyl mixture Aroclor 1254. In each situation, the sum of the levels of these 8 P-450 was at least as high as the spectrally determined P-450 content. Individual forms of P-450 can be induced by different compounds, and a single compound can lower the level of one form of P-450 while inducing one or more other forms of P-450. Catalytic activities toward each of the substrates observed with microsomal preparations are compared to rates predicted on the basis of the content of each of the 8 P-450. These studies provide a basis for further studies on the regulation of individual P-450, the physical properties of the different P-450 and the metabolic consequences of changes in the forms of P-450 in rat liver models.