Toward a systems approach to cytochrome P450 ensemble: interactions of CYP2E1 with other P450 species and their impact on CYP1A2
Toward a systems approach to cytochrome P450 ensemble: interactions of CYP2E1 with other P450 species and their impact on CYP1A2
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DOI：
10.1042/bcj20190532
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发表时间：
2019-12-01
影响因子：
4.1
通讯作者：
Davydov, Dmitri R.
中科院分区：
文献类型：
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作者：
Davydova, Nadezhda Y.;Dangi, Bikash;Davydov, Dmitri R.
在这项研究中，我们研究了乙醇诱导的CYP2E1与其他细胞色素P450物种相互作用并影响其底物代谢的能力。作为模型系统，我们使用了通过掺入纯化的CYP2E1获得的富含CYP2E1的人肝微粒体（HLM）。利用基于同源fret技术的CYP2E1低聚物标记BODIPY 577/618马来酰亚胺，我们证明了CYP2E1与HLM的相互作用导致其与存在于微粒体膜上的其他P450物种的混合低聚物的形成。CYP2E1的掺入导致CYP2E1特异性底物对硝基苯酚和氯虫臭氧的代谢率增加数倍。当蛋白质含量达到0.3-0.4 nmol/mg（或P450池中CYP2E1含量达到50%）时，其氧化速率与CYP2E1的掺入量成正比。合并的CYP2E1成为P450整体的全功能成员，与内源性CYP2E1没有任何可检测的功能差异。CYP2E1富集HLM导致7-乙氧基-4-氰香豆素（CEC）代谢的显著变化，CEC是CYP2C19和CYP1A2的底物，表明后者参与其代谢的程度增加。这种效应伴随着cyp1a2特异性底物7-乙氧基间苯磺酸脱烷基速率的增加。此外，通过探测CYP2E1与含有单个P450酶的模型微粒体的相互作用，我们发现CYP2E1与CYP1A2有效相互作用，但缺乏与CYP2C19形成复合物的能力。这一发现与cyp2e1诱导的CEC代谢主要途径从CYP2C19重定向到CYP1A2一致。
In this study, we investigate the ability of ethanol-inducible CYP2E1 to interact with other cytochrome P450 species and affect the metabolism of their substrates. As a model system, we used CYP2E1-enriched human liver microsomes (HLM) obtained by the incorporation of purified CYP2E1. Using a technique based on homo-FRET in oligomers of CYP2E1 labeled with BODIPY 577/618 maleimide we demonstrated that the interactions of CYP2E1 with HLM result in the formation of its mixed oligomers with other P450 species present in the microsomal membrane. Incorporation of CYP2E1 results in a multifold increase in the rate of metabolism of CYP2E1-specific substrates p-Nitrophenol and Chlorzaxozone. The rate of their oxidation remains proportional to the amount of incorporated CYP2E1 up to the content of 0.3-0.4 nmol/mg protein (or similar to 50% CYP2E1 in the P450 pool). The incorporated CYP2E1 becomes a fully functional member of the P450 ensemble and do not exhibit any detectable functional differences with the endogenous CYP2E1. Enrichment of HLM with CYP2E1 results in pronounced changes in the metabolism of 7-ethoxy-4-cyanocoumarin (CEC), the substrate of CYP2C19 and CYP1A2 suggesting an increase in the involvement of the latter in its metabolism. This effect goes together with an augmentation of the rate of dealkylation of CYP1A2-specific substrate 7-ethoxyresorufin. Furthermore, probing the interactions of CYP2E1 with model microsomes containing individual P450 enzymes we found that CYP2E1 efficiently interacts with CYP1A2, but lacks any ability to form complexes with CYP2C19. This finding goes inline with CYP2E1-induced redirection of the main route of CEC metabolism from CYP2C19 to CYP1A2.