Midazolam as a Probe for Drug-Drug Interactions Mediated by CYP3A4: Homotropic Allosteric Mechanism of Site-Specific Hydroxylation.
Midazolam as a Probe for Drug-Drug Interactions Mediated by CYP3A4: Homotropic Allosteric Mechanism of Site-Specific Hydroxylation.
复制标题
DOI：
10.1021/acs.biochem.1c00161
复制
发表时间：
2021-06-01
期刊：
影响因子：
2.9
通讯作者：
Sligar SG
中科院分区：
文献类型：
--
作者：
Denisov IG;Grinkova YV;Camp T;McLean MA;Sligar SG
我们利用咪达唑仑（MDZ）作为探针底物，开发了一种高效灵敏的检测人CYP3A4介导的药物-药物相互作用的探针。通过对几个实验数据集的四个参数进行全局分析，我们证明了第一个MDZ分子（MDZ1）在血红素铁附近的生产位点具有高亲和力，并且只在1位（1OH）发生羟基化。第二个咪达唑仑分子（MDZ2）结合在膜表面的变构位点，扰乱MDZ1的位置和迁移性，使4位的次要羟基化产物（4OH）以1:2的比例（35%）形成。在第二次MDZ结合后，没有观察到催化速率的增加。因此，1OH:4OH代谢比位点是药物（如黄体酮）的敏感探针，黄体酮与变构位点具有高亲和力结合并起效应器作用。我们观察到在黄体酮（PGS）存在下MDZ 1OH:4OH比率的类似变化，表明活性位点和变构位点之间存在直接通信。引入F-F '环的突变表明，残基F213和D214直接参与了导致MDZ同向协同性的变构相互作用，这些残基与L211一起参与了异向变构相互作用，其中PGS是效应体，MDZ是底物。分子动力学模拟为这种协作性的起源提供了一幅机制图。这些结果表明咪达唑仑可以作为人类P450 CYP3A4药物相互作用的敏感探针。
We developed an efficient and sensitive probe for drug–drug interactions mediated by human CYP3A4 by using midazolam (MDZ) as a probe substrate. Using global analysis of four parameters over several experimental data sets, we demonstrate that the first MDZ molecule (MDZ1) binds with high affinity at the productive site near the heme iron and gives only hydroxylation at the 1 position (1OH). The second midazolam molecule (MDZ2) binds at an allosteric site at the membrane surface and perturbs the position and mobility of MDZ1 such that the minor hydroxylation product at the 4 position (4OH) is formed in a 1:2 ratio (35%). No increase in catalytic rate is observed after the second MDZ binding. Hence, the site of the 1OH:4OH metabolism ratio is a sensitive probe for drugs, such as progesterone, that bind with high affinity to the allosteric site and serve as effectors. We observe similar changes in the MDZ 1OH:4OH ratio in the presence of progesterone (PGS), suggesting a direct communication between the active and allosteric sites. Mutations introduced into the F–F′ loop indicate that residues F213 and D214 are directly involved in allosteric interactions leading to MDZ homotropic cooperativity, and these same residues, together with L211, are involved in heterotropic allosteric interactions in which PGS is the effector and MDZ the substrate. Molecular dynamics simulations provide a mechanistic picture of the origin of this cooperativity. These results show that the midazolam can be used as a sensitive probe for drug–drug interactions in human P450 CYP3A4.
登录
查看更多内容
影响因子：
2.9
作者：
Hosea, NA;Miller, GP;Guengerich, FP
通讯作者：
Guengerich, FP
影响因子：
2.9
作者：
Domanski, TL;He, YA;Halpert, JR
通讯作者：
Halpert, JR
影响因子：
2
作者：
He, Fang;Zhang, Wen;Liu, Mingyi
通讯作者：
Liu, Mingyi
影响因子：
4.8
作者：
Frank, Daniel J.;Denisov, Ilia G.;Sligar, Stephen G.
通讯作者：
Sligar, Stephen G.
细胞色素P450的合作特性。
影响因子：
13.5
作者：
Denisov IG;Frank DJ;Sligar SG
通讯作者：
Sligar SG