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

基本信息

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
项目类别：
财政年份：
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

项目摘要

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（MOR）的选择性激动剂。在新型的合成阿片类药物（NSO）中，它们主导了休闲药市场，是阿片类药物危机的主要罪魁祸首，这已受到COVID-19的大流行而加剧。通过利用MOR的晶体结构，几个组研究了芬太尼的结合机制，但就结合方向和芬太尼构象而言，尚未达成共识。因此，芬太尼在MOR上的结合机制仍然是一个未解决且充满挑战的问题。在这里，我们进行了一项系统的计算研究，以研究首选的芬太尼构象，以及如何在MOR结合口袋中容纳这些构象。我们用元动力学模拟表征了芬太尼构象的自由能格局，并与长时间的分子动力学模拟进行了比较并评估了MOR中的几种可能的芬太尼结合条件。我们的结果表明，在没有受体的情况下，MOR结合口袋中最优选的结合姿势与芬太尼能量景观的全局最小值很好，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能会在替代结合姿势中稳定略微不满的芬太尼构象。通过将类似的研究扩展到芬太尼类似物，我们的发现在MOR上建立了芬太尼结合的结构活性关系。除了提供结构性基础以了解新兴NSO的潜在毒性外，这些见解还将有助于发展新的，更安全的镇痛药。 -Arrestin结合GPCR，以终止G蛋白信号传导并促进其他下游信号通路。使用单分子荧光共振能量转移成像，我们表明-arrestin在其基础状态下强烈自身抑制。它与模仿磷酸化受体尾巴的磷酸肽的互动有效地释放出-Arrestin尾巴从其N结构域释放，以采用明显的构象。出乎意料的是，我们发现与磷酸化受体结合的-Arrestin结合，具有与分离的磷酸肽相同的磷酸化条形码，高效效率高，并且需要激动剂促进的受体激活来激活-Arrestin激活，与续集受体C尾部的释放一致。这些发现以及聚焦的细胞研究表明，激动和受体C -tail释放是磷酸化受体激活-arrestin激活速率和效率的特定决定因素。我们推断该受体磷酸化模式与受体激动剂结合结合，协同建立了多种多样的下游-artrestin介导的事件的强度和特异性。