Activation of the food-derived mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine by rabbit and human liver microsomes and purified forms of cytochrome P-450.
Activation of the food-derived mutagen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine by rabbit and human liver microsomes and purified forms of cytochrome P-450.
复制标题
兔和人肝微粒体以及纯化形式的细胞色素 P-450 激活食品诱变剂 2-氨基-1-甲基-6-苯基咪唑并[4,5-b]吡啶。
作者：
M. McManus;J. Felton;M. Knize;W. Burgess;S. Roberts;S. Pond;I. Stupans;M. E. Veronese
使用对照和诱导的兔肝和肺微粒体以及6种纯化形式的细胞色素P-450，评估了2-氨基-1-甲基-6-苯基咪唑[4，5-B]吡啶（PhIP）致突变激活中涉及的兔细胞色素P-450的特异性。对照兔肝脏产生的回复突变体数量/2.5 μ g PhIP为260 +/- 196/10 μ g微粒体蛋白（平均值+/- SD; n = 3），当在艾姆斯试验中使用2，3，7，8-四氯二苯并-对-二恶英（TCDD）诱导的肝微粒体作为激活源时，这一数量增加至1265 +/- 248。微粒体从苯巴比妥，利福平和丙酮预处理的兔子没有增加的活动比对照。对照肺微粒体未将PhIP激活为诱变剂，而TCDD诱导的肺微粒体产生1443 +/- 136（平均值+/- SD; n = 4）艾姆斯/沙门氏菌回复突变体/100微克蛋白。在复溶实验中，发现细胞色素P450 4型和6型是PhIP对诱变剂的有效激活剂。在艾姆斯试验中，6型的活性是4型的3.1倍，在20分钟预孵育步骤中产生4577个回复突变体/10 pmol。细胞色素5型产生17个回复突变体/10 pmol，2型、3b型和3c型在将PhIP代谢为诱变剂方面没有活性。对照和诱导肝微粒体激活PhIP致突变剂的能力与其细胞色素P-450 4型（r = 0.97，r2 = 0.94）和6型（r = 0.95，r2 = 0.90）含量之间存在高度显著的统计学相关性。这些数据强烈支持参与多环烃诱导形式的细胞色素P450在激活的PhIP在兔。抗兔4型和6型IgG在7个具有可比mol. wt至兔细胞色素P-450形式4和6。然而，这些蛋白质的含量和人肝微粒体激活PhIP的能力之间不存在相关性。
The specificity of rabbit cytochrome P-450 involved in the mutagenic activation of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) was assessed using control and induced rabbit liver and lung microsomes, and six purified forms of cytochrome P-450. The number of revertants produced/2.5 micrograms PhIP by control rabbit liver was 260 +/- 196/10 micrograms of microsomal protein (mean +/- SD; n = 3), and this increased to 1265 +/- 248 when 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-induced liver microsomes were used as the activation source in the Ames test. Microsomes form phenobarbital-, rifampicin- and acetone-pretreated rabbits showed no increase in activity over controls. Control lung microsomes did not activate PhIP to a mutagen, whereas TCDD-induced lung microsomes produced 1443 +/- 136 (mean +/- SD; n = 4) Ames/Salmonella revertants/100 micrograms protein. In reconstitution experiments cytochrome P450 forms 4 and 6 were found to be efficient activators of PhIP to a mutagen Form 6 was 3.1-fold more active than form 4 and produced 4577 revertants/10 pmol with a 20-min preincubation step in the Ames test. Cytochrome form 5 produced 17 revertants/10 pmol and forms 2, 3b and 3c were not active in metabolizing PhIP to a mutagen. A highly significant statistical correlation existed between the capacity of control and induced liver microsomes to activate PhIP to a mutagen and their cytochrome P-450 form 4 (r = 0.97, r2 = 0.94) and form 6 (r = 0.95, r2 = 0.90) content. These data strongly support the involvement of polycyclic hydrocarbon-inducible forms of cytochrome P450 in the activation of PhIP in the rabbit. Anti-rabbit forms 4 and 6 IgGs recognized proteins in seven human liver microsomes of comparable mol. wt to rabbit cytochrome P-450 forms 4 and 6. However, no correlation existed between the content of these proteins and the capacity of human liver microsomes to activate PhIP.