Genetic Regulation of Aryl Hydrocarbon Hydroxylase Induction: V. SPECIFIC CHANGES IN SPIN STATE OF CYTOCHROME P<sub>450</sub> FROM GENETICALLY RESPONSIVE ANIMALS

The genetically controlled expression of hepatic aryl hydrocarbon (benzo[a]pyrene) hydroxylase induction in inbred and hybrid mice by aromatic hydrocarbons is associated, in freshly prepared oxidized microsomes, with increases in high spin cytochrome P450having a set of electron paramagnetic resonance signals at g = 8.0, 3.7, and 1.7. Phenobarbital causes similar rises (2- to 3-fold) in both high (g = 8.0, 3.7, and 1.7) and low spin (g = 2.4, 2.3, and 1.9) hepatic P450from any of the inbred or hybrid mice examined. Acetone treatment of microsomes destroys the g = 8.0, 3.7, 1.7 signals without affecting the hydroxylase activity, whereas Triton X-100 or sodium dodecyl sulfate affects the enzyme activity more so than the high spin signals. Substrate binding produces interconversion between high and low spin P450. treatment of microsomes with sodium dodecyl sulfate or sodium deoxycholate or at 55° for 5 min causes the irreversible loss of high and low spin signals concomitant with a rise in the g = 5.95 signal. Induction of the hydroxylase activity is associated with the increased g = 8.0, 3.7, and 1.7 signal heights in the kidney from 3-methylcholanthrene-treated rabbit and in the liver or kidney from 3-methylcholanthrene-treated rat, but not in liver from aromatic hydrocarbon-treated rabbit. Hence, aryl hydrocarbon hydroxylase induction by polycyclic hydrocarbons and the increase in high-spin cytochrome P450are not always necessarily related.These results indicate that aromatic hydrocarbon treatment of the genetically responsive animal, rather than treatment of any animal with aromatic hydrocarbons , causes an increase in a P450species which has iron in a preferred high spin configuration. This preferred spin state may reflect a change near the P450active site, such as the presence of new apoenzyme(s) or a different configuration. There is another high spin protein iron having a g = 5.95 signal; this most likely represents a denatured form of P450.

芳香烃对近交系和杂交小鼠肝脏芳基烃（苯并[a]芘）羟化酶诱导的基因控制表达，在新鲜制备的氧化微粒体中，与高自旋细胞色素P450的增加有关，其在g = 8.0、3.7和1.7处有一组电子顺磁共振信号。苯巴比妥使所检测的任何近交系或杂交小鼠肝脏中高自旋（g = 8.0、3.7和1.7）和低自旋（g = 2.4、2.3和1.9）的P450都出现类似的升高（2 - 3倍）。丙酮处理微粒体会破坏g = 8.0、3.7、1.7的信号，但不影响羟化酶活性，而Triton X - 100或十二烷基硫酸钠对酶活性的影响比对高自旋信号的影响更大。底物结合导致高自旋和低自旋P450之间的相互转换。用十二烷基硫酸钠或脱氧胆酸钠处理微粒体，或在55°C处理5分钟，会导致高自旋和低自旋信号不可逆地丧失，同时g = 5.95信号升高。羟化酶活性的诱导与3 - 甲基胆蒽处理的兔子肾脏以及3 - 甲基胆蒽处理的大鼠肝脏或肾脏中g = 8.0、3.7和1.7信号高度的增加有关，但在芳香烃处理的兔子肝脏中无关。因此，多环烃对芳基烃羟化酶的诱导和高自旋细胞色素P450的增加并不总是必然相关。这些结果表明，对具有遗传响应的动物进行芳香烃处理，而不是对任何动物进行芳香烃处理，会导致一种P450物种的增加，该物种中的铁处于优选的高自旋构型。这种优选的自旋状态可能反映了P450活性位点附近的变化，例如新的脱辅基酶的存在或不同的构型。还有另一种高自旋蛋白铁，其具有g = 5.95的信号；这很可能代表P450的一种变性形式。