Molecular Basis for Benzodiazepine Agonist Action at the Type 1 Cholecystokinin Receptor
Molecular Basis for Benzodiazepine Agonist Action at the Type 1 Cholecystokinin Receptor
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DOI：
10.1074/jbc.m113.480715
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发表时间：
2013-07-19
影响因子：
4.8
通讯作者：
Miller, Laurence J.
中科院分区：
文献类型：
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作者：
Harikumar, Kaleeckal G.;Cawston, Erin E.;Miller, Laurence J.
了解药物作用的分子基础有助于开发更有效和更具选择性的药物。在这里，我们探索一种独特的小分子配体的作用分子基础，它是一种1型CCK受体激动剂和2型CCK受体拮抗剂GI181771X。我们利用结构相关的放射性碘化配体选择性地结合使用相同变构配体结合口袋的CCK受体亚型，使用野生型受体和嵌合结构交换该口袋内不同的残基来表征其结合。测定细胞内钙离子浓度以确定其生物学活性。小分子激动剂对接到1型CCK受体的分子模型是用配体引导的精制方法开发的。最优模型不同于以前针对同一受体的拮抗剂模型，并且与当前的突变数据在力学上是一致的。这项研究揭示了亮氨酸(7.39)的一个关键作用，该作用被预测为与苯二氮卓类药物N1位的异丙基相互作用，苯二氮卓是生物活性的“触发器”。该分子模型预测了其他小分子激动剂的结合，有效地将它们与1065个经批准的药物诱饵区分开来，曲线下面积为99%。该模型还选择性地丰富了激动剂化合物，当播种在1型CCK受体的2175个非激动剂配体(曲线下面积78%)时，ROC分析确定了130种激动剂。这一系列中的苯二氮卓类激动剂以一致的姿势停靠在这个口袋里，其中Leu7.39起着关键作用，而对于化学上不同的激动剂来说，这个残留物的作用不太清楚。
Understanding the molecular basis of drug action can facilitate development of more potent and selective drugs. Here, we explore the molecular basis for action of a unique small molecule ligand that is a type 1 cholecystokinin (CCK) receptor agonist and type 2 CCK receptor antagonist, GI181771X. We characterize its binding utilizing structurally related radioiodinated ligands selective for CCK receptor subtypes that utilize the same allosteric ligand-binding pocket, using wild-type receptors and chimeric constructs exchanging the distinct residues lining this pocket. Intracellular calcium assays were performed to determine biological activity. Molecular models for docking small molecule agonists to the type 1 CCK receptor were developed using a ligand-guided refinement approach. The optimal model was distinct from the previous antagonist model for the same receptor and was mechanistically consistent with the current mutagenesis data. This study revealed a key role for Leu(7.39) that was predicted to interact with the isopropyl group in the N1 position of the benzodiazepine that acts as a "trigger" for biological activity. The molecular model was predictive of binding of other small molecule agonists, effectively distinguishing these from 1065 approved drug decoys with an area under curve value of 99%. The model also selectively enriched for agonist compounds, with 130 agonists identified by ROC analysis when seeded in 2175 non-agonist ligands of the type 1 CCK receptor (area under curve 78%). Benzodiazepine agonists in this series docked in consistent pose within this pocket, with a key role played by Leu7.39, whereas the role of this residue was less clear for chemically distinct agonists.