Vanderslice R R, Domin B A, Carver G T, Philpot R M
Mol Pharmacol. 1987 Apr;31(4):320-5.
The presence of homologues of rabbit cytochrome P-450 isozyme 5 in pulmonary and hepatic microsomal preparations from guinea pig, mouse, monkey, hamster, and rat was examined by immunoblotting and inhibition of metabolism of 2-aminofluorene with antibodies to isozyme 5. Homologues to isozyme 5 were detected in pulmonary preparations from all five species. However, only hepatic preparations from hamster, in addition to those from rabbit, contained detectable levels of this isozyme. With the exception of induction by phenobarbital in rabbit liver, treatment of animals with phenobarbital or tetrachlorodibenzo-p-dioxin did not increase hepatic or pulmonary content of isozyme 5 homologues or the amount of 2-aminofluorene metabolism inhibited by antibodies to isozyme 5. Metabolism of 2-aminofluorene was measured both colorimetrically (formation of a reduced iron chelate from the N-hydroxyfluorene metabolite) and radiochemically (separation of 3H-metabolites by high performance liquid chromatography and quantitation by scintillation counting). A turnover number of 48 nmol of product X min-1 X nmol of enzyme-1 for isozyme 5-catalyzed metabolism of 2-aminofluorene was determined with incubations containing isozyme 5 purified from rabbit lung. A similar turnover number was calculated from the rabbit hepatic microsomal activity inhibited by antibodies to isozyme 5 and the microsomal isozyme 5 content measured by immunoquantitation. In other species, amounts of metabolism inhibited by antibodies to isozyme 5 agreed qualitatively with relative staining intensities on immunoblots. In all species except the hamster, rates of total and isozyme 5-catalyzed metabolism of 2-aminofluorene were greater with pulmonary than with hepatic microsomal preparations from untreated animals.
通过免疫印迹法以及用针对同工酶5的抗体抑制2-氨基芴的代谢，检测了豚鼠、小鼠、猴子、仓鼠和大鼠的肺和肝微粒体制剂中兔细胞色素P-450同工酶5同源物的存在情况。在所有五个物种的肺制剂中均检测到了同工酶5的同源物。然而，除了兔的肝制剂外，只有仓鼠的肝制剂含有可检测水平的这种同工酶。除了苯巴比妥诱导兔肝中的情况外，用苯巴比妥或四氯二苯并对二恶英处理动物并没有增加同工酶5同源物的肝或肺含量，也没有增加被针对同工酶5的抗体抑制的2-氨基芴代谢量。2-氨基芴的代谢通过比色法（由N-羟基芴代谢物形成还原铁螯合物）和放射化学法（通过高效液相色谱法分离3H-代谢物并通过闪烁计数进行定量）进行测定。用从兔肺中纯化的同工酶5进行孵育，测定了同工酶5催化2-氨基芴代谢的转换数为48 nmol产物·min⁻¹·nmol酶⁻¹。根据被针对同工酶其5的抗体抑制的兔肝微粒体活性以及通过免疫定量测定的微粒体同工酶5含量，计算出了类似的转换数。在其他物种中，被针对同工酶5的抗体抑制的代谢量在质量上与免疫印迹上的相对染色强度一致。在除仓鼠外的所有物种中，未处理动物的肺微粒体制剂中2-氨基芴的总代谢率和同工酶5催化的代谢率均高于肝微粒体制剂。
