Department of Pharmacology, University of Michigan, Ann Arbor, MI 48109-5632, USA.
Drug Metab Dispos. 2010 Dec;38(12):2286-92. doi: 10.1124/dmd.110.034710. Epub 2010 Sep 8.
Although the ability of disulfiram to inactivate CYP2E1 has been known for more than 20 years, the mechanism has not yet been elucidated. A metabolite of disulfiram, diethyldithocarbamate (DDC), is converted by CYP2E1 to a reactive intermediate that subsequently inactivates the protein, leading to mechanism-based inactivation. Mass spectral analysis of the inactivated human 2E1 protein demonstrates that the inactivation is due to the formation of an adduct of the reactive metabolite of DDC with the apoprotein. These data, along with mass spectral analysis of a reactive intermediate trapped with GSH, indicate the involvement of a reactive intermediate with a molecular mass of 116 Da. Our results suggest that this binding involves formation of a disulfide bond with one of the eight cysteines in CYP2E1. The inactivation of wild-type CYP2E1 as well as two of its polymorphic mutants, CYP2E12 and CYP2E14, was also investigated. For wild-type CYP2E1, the K(I) was 12.2 μM and the k(inact) was 0.02 min(-1). The K(I) values for the two polymorphic mutants were 227.6 and 12.4 μM for CYP2E1.2 and CYP2E1.4, and the k(inact) values were 0.0061 and 0.0187 min(-1), respectively. These data indicate that DDC is a much less efficient inactivator of CYP2E1.2 than it is of either the wild-type or the CYP2E1.4 variant.
尽管双硫仑使 CYP2E1 失活的能力已经为人所知超过 20 年，但该机制尚未阐明。双硫仑的一种代谢物，二乙基二硫代氨基甲酸盐（DDC），被 CYP2E1 转化为一种反应性中间产物，随后使该蛋白失活，导致基于机制的失活。对失活的人 2E1 蛋白的质谱分析表明，失活是由于 DDC 的反应性代谢物与脱辅基蛋白形成加合物所致。这些数据，以及用 GSH 捕获的反应性中间产物的质谱分析，表明涉及分子量为 116 Da 的反应性中间产物。我们的研究结果表明，这种结合涉及与 CYP2E1 中的八个半胱氨酸之一形成二硫键。还研究了野生型 CYP2E1 以及其两种多态性突变体 CYP2E12 和 CYP2E14 的失活情况。对于野生型 CYP2E1，K(I)为 12.2 μM，k(inact)为 0.02 min(-1)。对于两种多态性突变体 CYP2E1.2 和 CYP2E1.4，K(I)值分别为 227.6 和 12.4 μM，k(inact)值分别为 0.0061 和 0.0187 min(-1)。这些数据表明，DDC 是 CYP2E1.2 的失活剂，其效率远低于野生型或 CYP2E1.4 变体。
