细胞色素 P450 3A4 在底物和细胞色素 P450 还原酶存在下的结构动力学,Biochemistry

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细胞色素 P450 3A4 在底物和细胞色素 P450 还原酶存在下的结构动力学
Biochemistry ( IF 2.9 ) Pub Date : 2021-07-01 , DOI: 10.1021/acs.biochem.1c00178
Julie Ducharme ₁ , Irina F Sevrioukova ₂ , Christopher J Thibodeaux ₁ , Karine Auclair ₁

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细胞色素 P450 3A4 (CYP3A4) 是人类最重要的药物代谢酶，与有害的药物相互作用有关。已知 CYP3A4 的活性受几种化合物和电子转移伴侣细胞色素 P450 还原酶 (CPR) 的调节。然而，人们对这些影响的潜在机制知之甚少。我们使用氢-氘交换质谱来研究 CPR 和三种不同底物（7-苄氧基-4-三氟甲基-香豆素、睾酮和孕酮）的结合对 CYP3A4 构象动力学的影响。在这里，我们报告 CYP3A4 与底物或与 CPR 的氧化或还原形式的相互作用导致 CYP3A4 结构的整体僵化。这从 CYP3A4 的几个区域的氘交换抑制中可以看出，包括已知参与蛋白质-蛋白质相互作用（螺旋 C）和底物结合和特异性（螺旋 B' 和 E，以及环 K/β1）的区域。此外，观察到的一些 CYP3A4 衍生肽的双峰同位素分布在与 CPR 和/或底物结合时受到极大影响，表明存在受配体/CPR 结合干扰的稳定 CYP3A4 构象群。该结果对理解 CYP 酶中配体结合、变构和催化的机制具有重要意义。表明存在受配体/CPR结合干扰的稳定CYP3A4构象群。该结果对理解 CYP 酶中配体结合、变构和催化的机制具有重要意义。表明存在受配体/CPR结合干扰的稳定CYP3A4构象群。该结果对理解 CYP 酶中配体结合、变构和催化的机制具有重要意义。

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Structural Dynamics of Cytochrome P450 3A4 in the Presence of Substrates and Cytochrome P450 Reductase

Cytochrome P450 3A4 (CYP3A4) is the most important drug-metabolizing enzyme in humans and has been associated with harmful drug interactions. The activity of CYP3A4 is known to be modulated by several compounds and by the electron transfer partner, cytochrome P450 reductase (CPR). The underlying mechanism of these effects, however, is poorly understood. We have used hydrogen–deuterium exchange mass spectrometry to investigate the impact of binding of CPR and of three different substrates (7-benzyloxy-4-trifluoromethyl-coumarin, testosterone, and progesterone) on the conformational dynamics of CYP3A4. Here, we report that interaction of CYP3A4 with substrates or with the oxidized or reduced forms of CPR leads to a global rigidification of the CYP3A4 structure. This was evident from the suppression of deuterium exchange in several regions of CYP3A4, including regions known to be involved in protein–protein interactions (helix C) and substrate binding and specificity (helices B′ and E, and loop K/β1). Furthermore, the bimodal isotopic distributions observed for some CYP3A4-derived peptides were drastically impacted upon binding to CPR and/or substrates, suggesting the existence of stable CYP3A4 conformational populations that are perturbed by ligand/CPR binding. The results have implications for understanding the mechanisms of ligand binding, allostery, and catalysis in CYP enzymes.

更新日期：2021-07-20

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