Structure and Function of a Serotonin Transporter

血清素转运蛋白的结构和功能

基本信息

批准号：
8773997
负责人：
SATINDER Kaur SINGH
金额：
$ 57.87万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-08-15 至 2019-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8773997
关键词：
2-cyclopentyl-5-(5-isoquinolylsulfonyl)-6-nitro-1H-benzo(D)imidazole Adverse effects Affinity Amphetamines Antidepressive Agents Anxiety Applications Grants Attention Autistic Disorder Binding Binding Sites Cells Chemicals Clinic Clinical Cocaine Complex Coupled Coupling Cysteine Cytoplasm Data Detergents Deuterium Disease Dissociation Drosophila melanogaster Drug Design Drug resistance Exhibits Functional disorder Genetic Polymorphism Goals Homologous Gene Human Hydrogen Investigation Ion Cotransport Ions Kinetics Laboratories Ligands Light Liposomes Mass Spectrum Analysis Measures Membrane Mental Depression Modeling Molecular Molecular Conformation Molecular Machines Movement Neurons Obsessive-Compulsive Disorder Orthologous Gene Paroxetine Pharmacology Phospholipids Play Preparation Process Proteins Psychotropic Drugs Regulation Role Selective Serotonin Reuptake Inhibitor Serotonin Shapes Signal Transduction Site-Directed Mutagenesis Structure Synapses Synaptic Transmission Testing Therapeutic Vestibule Work X-Ray Crystallography antiport base crosslink design ecstasy extracellular in vivo inhibitor/antagonist insight nanodisk neuropsychiatry neurotransmission public health relevance reconstitution research study serotonin transporter success

项目摘要

DESCRIPTION (provided by applicant): The plasmalemmal serotonin transporter (SERT) terminates serotonergic neurotransmission and thus plays a critical role in shaping the duration and magnitude of synaptic signaling. It works by coupling preexisting ion gradients to the thermodynamically unfavorable movement of serotonin from the synapse to neuronal and glial cytoplasms. This molecular machine has garnered significant clinical attention because its dysfunction has been implicated in multiple debilitating neuropsychiatric diseases such as depression, autism, obsessive- compulsive disorder, and generalized anxiety. Furthermore, it is the target of numerous psychoactive agents such as antidepressants, cocaine, and the amphetamine derivative "ecstasy". Despite such clinical and pharmacological significance, atomic-level detail into the mechanism of substrate translocation, inhibition, and regulation has remained elusive. Two goals of this proposal include 1) solving the structure of a SERT in complex with substrates, ions, and inhibitors; and 2) probing the conformational dynamics of these interactions via hydrogen-deuterium exchange mass spectrometry (HDX-MS). Structure-based hypotheses will subsequently be tested with a combination of site-directed mutagenesis, cysteine crosslinking, transient and steady-state flux kinetics, dissociation/association binding kinetics, X-ray crystallography, and HDX-MS. Success will permit further investigation into specific antagonist binding sites and perhaps into the molecular basis for drug resistance, thereby paving the way for rational drug design efforts. When eventually coupled with in vivo work beyond the scope of this proposal, SERT structure/function studies may also shed light on the molecular underpinnings of disease-associated polymorphisms. Achieving any of one of these objectives would likely have significant impact in both the laboratory and the clinic.

描述（由申请人提供）：血浆5-羟色胺转运蛋白（SERT）终止血清素能神经传递，因此在塑造突触信号的持续时间和幅度中起着至关重要的作用。它通过将先前存在的离子梯度与5-羟色胺从突触到神经元和神经胶质细胞肿瘤的热力学不利运动耦合而起作用。这款分子机器人引起了极大的临床注意力，因为其功能障碍与多种使人衰弱的神经精神疾病有关，例如抑郁症，自闭症，强迫症和普遍焦虑。此外，它是众多精神活性剂的靶标，例如抗抑郁药，可卡因和苯丙胺衍生物“摇头丸”。尽管如此临床和药理意义，但原子水平的细节涉及底物易位，抑制和调节的机制仍然难以捉摸。该提案的两个目标包括1）解决与底物，离子和抑制剂复杂的SERT结构； 2）通过氢欧交换质谱法（HDX-MS）探测这些相互作用的构象动力学。基于结构的假设随后将通过位置定向诱变，半胱氨酸交联，瞬态和稳态通量动力学，分离/关联结合动力学，X射线晶体学和HDX-MS的组合进行测试。成功将允许进一步研究特定的拮抗剂结合位点，并可能成为耐药性的分子基础，从而为理性的药物设计工作铺平了道路。当最终与该提案范围之外的体内工作结合在一起时，SERT结构/功能研究也可能揭示出与疾病相关的多态性的分子基础。实现这些目标中的任何一个都可能对实验室和诊所产生重大影响。