Structural basis for the functions of dopamine receptors, neurotransmitter transporters, and sigma 1 receptor

多巴胺受体、神经递质转运蛋白和 sigma 1 受体功能的结构基础

• 批准号: 10699660
• 负责人: Lei Shi
• 金额: $ 218.05万
• 依托单位: NATIONAL INSTITUTE ON DRUG ABUSE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10699660
• 关键词: Agonist; Amphetamines; Analgesics; Arrestins; Bar Codes; Binding; Binding Sites; Biological Models; Brain; COVID-19 pandemic; Carrier Proteins; Characteristics; Cocaine; Consensus; Crystallization; Development; Dopamine Receptor; Drug Targeting; Event; Fentanyl; Free Energy; G-Protein-Coupled Receptors; GTP-Binding Proteins; Goals; Heroin; Human; Image; Investigation; Lead; Ligand Binding; Ligands; Mediating; Molecular Conformation; N Domain; Nervous System; Neurons; Neurotransmitters; Opioid Receptor; Opioid Receptor Binding; Overdose; Pattern; Pharmaceutical Preparations; Pharmacology; Pharmacotherapy; Phosphopeptides; Phosphorylation; Play; Proteins; Receptor Activation; Recreational Drugs; Research; Research Subjects; Role; Signal Pathway; Signal Transduction; Signaling Protein; Specificity; Structure; Structure-Activity Relationship; Substance Use Disorder; Tail; Therapeutic; Therapeutic Intervention; Toxic effect; addiction; analog; computer studies; conformational conversion; design; dopamine transporter; drug discovery; drug market; drug of abuse; extracellular; insight; molecular dynamics; nervous system disorder; novel; opioid epidemic; receptor; scaffold; serotonin transporter; sigma-1 receptor; simulation; single-molecule FRET; small molecule; synergism; synthetic cannabinoid; synthetic opioid; therapeutic development; therapeutically effective; therapy development

Fentanyl and its analogs are selective agonists of the micro-opioid receptor (MOR). Among novel synthetic opioids (NSOs), they dominate the recreational drug market and are the main culprits for the opioid crisis, which has been exacerbated by the COVID-19 pandemic. By taking advantage of the crystal structures of the MOR, several groups have investigated the binding mechanism of fentanyl, but have not reached a consensus, in terms of both the binding orientation and the fentanyl conformation. Thus, the binding mechanism of fentanyl at the MOR remains an unsolved and challenging question. Here, we carried out a systematic computational study to investigate the preferred fentanyl conformations, and how these conformations are being accommodated in the MOR binding pocket. We characterized the free energy landscape of fentanyl conformations with metadynamics simulations, and compared and evaluated several possible fentanyl binding conditions in the MOR with long-timescale molecular dynamics simulations. Our results indicate that the most preferred binding pose in the MOR binding pocket corresponds well with the global minimum on the energy landscape of fentanyl in the absence of the receptor, while the energy landscape can be reconfigured by modifying the fentanyl scaffold. The interactions with the receptor may stabilize a slightly unfavored fentanyl conformation in an alternative binding pose. By extending similar investigations to fentanyl analogs, our findings establish a structure-activity relationship of fentanyl binding at the MOR. In addition to providing a structural basis to understand the potential toxicity of the emerging NSOs, such insights will contribute to developing new, safer analgesics. -arrestins bind GPCRs to terminate G protein signaling and to facilitate other downstream signaling pathways. Using single-molecule fluorescence resonance energy transfer imaging, we show that -arrestin is strongly autoinhibited in its basal state. Its engagement with a phosphopeptide mimicking phosphorylated receptor tail efficiently releases the -arrestin tail from its N domain to assume distinct conformations. Unexpectedly, we find that -arrestin binding to phosphorylated receptor, with a phosphorylation barcode identical to the isolated phosphopeptide, is highly inefficient and that agonist-promoted receptor activation is required for -arrestin activation, consistent with the release of a sequestered receptor C tail. These findings, together with focused cellular investigations, reveal that agonism and receptor C-tail release are specific determinants of the rate and efficiency of -arrestin activation by phosphorylated receptor. We infer that receptor phosphorylation patterns, in combination with receptor agonism, synergistically establish the strength and specificity with which diverse, downstream -arrestin-mediated events are directed.

芬太尼及其类似物是微阿片受体（莫尔）的选择性激动剂。在新型合成阿片类药物（NSOs）中，它们主导着娱乐性药物市场，是阿片类药物危机的罪魁祸首，COVID-19大流行加剧了这一危机。通过利用莫尔的晶体结构，几个小组已经研究了芬太尼的结合机制，但在结合取向和芬太尼构象方面尚未达成共识。因此，芬太尼在莫尔的结合机制仍然是一个未解决的和具有挑战性的问题。在这里，我们进行了系统的计算研究，调查首选芬太尼构象，以及如何将这些构象被容纳在莫尔结合口袋。我们的特点是自由能景观的芬太尼构象与metadhesics模拟，并比较和评估了几种可能的芬太尼结合条件的莫尔与长时间尺度的分子动力学模拟。我们的研究结果表明，在莫尔结合口袋中的最优选的结合姿势与芬太尼在没有受体的情况下的能量景观上的全局最小值相对应，而能量景观可以通过修改芬太尼支架来重新配置。与受体的相互作用可以使稍微不利的芬太尼构象稳定在替代结合姿势中。通过对芬太尼类似物进行类似的研究，我们的研究结果建立了芬太尼在莫尔比上的结构-活性关系。除了为了解新出现的国家统计局的潜在毒性提供结构基础外，这种见解还将有助于开发新的、更安全的镇痛剂。 - 抑制蛋白结合GPCR以终止G蛋白信号传导并促进其他下游信号传导途径。使用单分子荧光共振能量转移成像，我们表明，-arrestin是强烈的自抑制在其基础状态。它与一个磷酸肽模仿磷酸化受体尾巴有效地释放-arrestin尾巴从其N结构域承担不同的构象。出乎意料的是，我们发现，-arrestin结合磷酸化受体，具有与分离的磷酸肽相同的磷酸化条形码，是非常低效的，并且激动剂促进的受体活化是-arrestin活化所需的，与隔离的受体C尾的释放一致。这些研究结果，再加上集中的细胞调查，揭示激动和受体C-尾释放的磷酸化受体的抑制蛋白激活的速率和效率的具体决定因素。我们推断，受体磷酸化模式，与受体激动作用相结合，协同建立的强度和特异性与不同的，下游抑制蛋白介导的事件是直接的。