Functional Mechanisms of NTs in a Structural Context

结构背景下 NT 的功能机制

• 批准号: 6880178
• 负责人: HAREL WEINSTEIN
• 金额: $ 18.75万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-12-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6880178
• 关键词: computer simulation; dopamine transporter; drug abuse; electron microscopy; membrane proteins; neurotransmitter transport; norepinephrine; phosphorylation; protein protein interaction; protein structure function; protein transport; serotonin transporter

This Project of the PPG is devoted to the structural context of the mechanistic study of Na+/Cl- dependent biogenic amine transporters (NT) function. We will investigate - for DAT, SERT, and NET - the role of specific structural elements in the physiological processes underlying neurotransmission and mechanisms in drug addiction and abuse, with generalizations and comparisons to other NTs. The proposed studies use 3D modeling and computational analysis of dynamics and energetics to achieve, in close collaboration with the other PPG projects, a quantitative evaluation of the structural and energetic basis for: (i) the role of specific sets of both intra- and inter-molecular interactions of N-termini, C-termini and key loops of NTs in experimentally measured properties that underlie function, and the changes that are produced by phosphorylation (SA #1a), and (ii) the detailed mode of interaction and specificity for interactions with PDZ domains considered to be involved in internalization, recycling, sorting, stabilization of NT, and in regulation of NT function. We will investigate steric and dynamic properties of NT C-terminal interaction with PDZ domains (starting with DAT-PICK1 interactions), and the molecular determinants for regulation of interactions selectivity by phosphorylation (of either the peptide or the PDZ) (SA #1b). The considerable amounts and variety of data prompt us to build two modes of empowering representation: a) 3D models based on structure-informed working hypotheses, refined bioinformatics approaches and computational molecular biophysics algorithms, constraints from functional insights obtained in the PPG and the literature, and newly published structural information from x-ray and EM studies of transporter and membrane protein systems that can be used as guides for structural principles in 3D modeling and as probes for validation of approaches (SA #2), and b) a transporter-centric information management system (TIMS) to serve as an electronic catalog of quantitative data about NT interactions (with ligands: substrates, blockers and modulators, and cell components such as scaffolding proteins), as well as NT activities (e.g., substrate transport rates under various conditions and NT states) and other quantitative data obtained in the PPG Projects (SA #3).

PPG 的这个项目致力于 Na+/Cl 依赖性生物胺转运蛋白 (NT) 功能的机制研究的结构背景。我们将研究 DAT、SERT 和 NET 的特定结构元素在神经传递的生理过程中的作用以及药物成瘾和滥用的机制，并与其他 NT 进行概括和比较。拟议的研究使用动力学和能量学的 3D 建模和计算分析，与其他 PPG 项目密切合作，对结构和能量基础进行定量评估：（i）NT 的 N 端、C 端和关键环的分子内和分子间相互作用的特定组的作用 实验测量了功能基础的特性以及磷酸化产生的变化 (SA #1a)，以及 (ii) 相互作用的详细模式以及与 PDZ 结构域相互作用的特异性，这些结构域被认为涉及 NT 的内化、回收、分类、稳定和 NT 功能的调节。我们将研究 NT C 末端与 PDZ 结构域相互作用的空间和动态特性（从 DAT-PICK1 相互作用开始），以及通过磷酸化（肽或 PDZ）调节相互作用选择性的分子决定因素 (SA #1b)。大量和多样化的数据促使我们建立两种赋权表示模式：a）基于结构信息工作假设的 3D 模型、精炼的生物信息学方法和计算分子生物物理学算法、PPG 和文献中获得的功能见解的约束以及新的模型。 发表了转运蛋白和膜蛋白系统的 X 射线和 EM 研究的结构信息，可用作 3D 建模中结构原理的指南和方法验证的探针（SA #2），以及 b）以转运蛋白为中心的信息管理系统（TIMS），作为有关 NT 相互作用的定量数据的电子目录（与配体：底物、阻断剂和调节剂以及细胞成分，例如 支架蛋白），以及 NT 活性（例如，各种条件和 NT 状态下的底物转运速率）以及 PPG 项目（SA #3）中获得的其他定量数据。