3-甲基吲哚甲基氧化特异性催化剂CYP4B2的表征。
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卡尔BA, Ramakanth S, Dannan GA, Yost GS
3-甲基吲哚甲基氧化特异性催化剂CYP4B2的表征。
Mol Pharmacol, 2003五月;63(5):1137-47。
- PubMed ID
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12695542 (PubMed视图］
- 摘要
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化学物质对肺组织的选择性毒性主要是由某些肺细胞色素P450酶的选择性表达介导的。本文描述了从山羊肺中提取的一种酶CYP4B2的纯化、克隆和鉴定。经多级离子交换色谱分离纯化的P450酶，电泳均匀，表观分子质量为55000 Da。Western blotting研究表明，CYP4B酶在兔、大鼠和小鼠的肺组织中有选择性表达。从山羊肺组织中克隆了CYP4B2和CYP4B2v两个cdna。CYP4B2被预测为511个氨基酸，与四种已知的CYP4B1蛋白约82%相似。同时，已知的人类CYP4B1 cDNA的一个变体，包含S207插入，从人类肺组织中克隆出来。经修饰的重组山羊CYP4B2在大肠杆菌中表达，酶催化原型底物2AF的n -羟基化。CYP4B2优先脱氢，而不是羟化，空气毒性3-甲基吲哚(3MI) (V(max)分别为4.61和0.83 nmol/nmol的P450/min)。为了研究共价血红素结合CYP4酶在CYP4B2介导的3MI代谢中的相关性，我们评估了一个位点定向突变体(CYP4B2/A315E)。 The mutation had little effect on the V(max) of either dehydrogenation or hydroxylation but increased the K(m), which decreased the catalytic efficiency (V/K) for 3MI. The A315E mutation shifted the absorbance maximum of the enzyme from 448 to 451 nm, suggesting that the electron density of the heme was altered. These results demonstrate that CYP4B2 is highly specific for methyl group oxidation of 3MI, without formation of ring-oxidized metabolites, and seems to be predominately responsible for the highly organ-specific toxicity of 3MI in goats.