Department of Medicinal Chemistry , University of Washington , Seattle , Washington 98195-7610 , United States.
Department of Physiology and Biophysics and the Massey Cancer Center, School of Medicine , Virginia Commonwealth University , Richmond , Virginia 23298-0035 , United States.
Biochemistry. 2020 Feb 18;59(6):766-779. doi: 10.1021/acs.biochem.9b01001. Epub 2020 Jan 27.
Promiscuous and allosteric drug interactions with cytochrome P450 3A4 (CYP3A4) are ubiquitous but incompletely understood at the molecular level. A classic allosteric CYP3A4 drug interaction includes the benzodiazepine midazolam (MDZ). MDZ exhibits homotropic and heterotropic allostery when metabolized to 1'-hydroxy and 4-hydroxy metabolites in varying ratios. The combination of hydrogen-deuterium exchange mass spectrometry (HDX-MS) and Gaussian accelerated molecular dynamics (GaMD) simulations of CYP3A4 in lipid nanodiscs and in a lipid bilayer, respectively, reveals MDZ-dependent changes in dynamics in a membrane environment. The F-, G-, and intervening helices, as well as the loop preceding the β1-sheets, display the largest observed changes in HDX. The GaMD suggests a potential allosteric binding site for MDZ in the F'- and G'-regions, which undergo significant increases in HDX at near-saturating MDZ concentrations. The HDX-MS and GaMD results confirm that changes in dynamics are most significant near the developing consensus allosteric site, and these changes are distinct from those observed previously with the nonallosteric inhibitor ketoconazole. The results suggest that the allosteric MDZ remains mobile in its binding site at the Phe-cluster. The results further suggest that this binding site remains dynamic or changes the depth of insertion in the membrane.
细胞色素 P450 3A4（CYP3A4）的混杂和变构药物相互作用普遍存在，但在分子水平上仍不完全了解。经典的变构 CYP3A4 药物相互作用包括苯并二氮䓬类药物咪达唑仑（MDZ）。MDZ 在代谢为 1'-羟基和 4-羟基代谢物时表现出同型变构和异型变构，其比例不同。分别使用氢氘交换质谱（HDX-MS）和高斯加速分子动力学（GaMD）模拟在脂质纳米盘中和脂质双层中的 CYP3A4，揭示了在膜环境中 MDZ 依赖性的动力学变化。F、G 和 intervening 螺旋以及β1 片层之前的环显示出 HDX 中观察到的最大变化。GaMD 表明 MDZ 在 F'和 G'区域可能存在变构结合位点，在接近饱和的 MDZ 浓度下，这些区域的 HDX 显著增加。HDX-MS 和 GaMD 结果证实，在正在形成的共识变构位点附近，动力学变化最为显著，这些变化与以前观察到的非变构抑制剂酮康唑的变化不同。结果表明，变构 MDZ 在其结合在 Phe 簇的位点上保持移动性。结果进一步表明，该结合位点保持动态或改变在膜中的插入深度。
