Allosteric Enhancement of Adenosine Receptors

腺苷受体的变构增强

• 批准号: 7340393
• 负责人: Joel M. Linden
• 金额: $ 36.78万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-04-20 至 2009-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7340393
• 关键词: Adenosine; Adenosine A1 Receptor; Adenosine A2A Receptor; Adverse effects; Affect; Affinity; Agonist; Alkylating Agents; Allosteric Site; Amino Acid Sequence; Animal Model; Animals; Aromatic Amines; Australia; Benzodiazepines; Binding; Binding Sites; Biological Assay; Blood - brain barrier anatomy; Boxing; Brain; Cardiac; Cations; Cells; Central Nervous System Diseases; Chemicals; Chemistry; Chimera organism; Class; Collaborations; Complex; Coupling; Cyclic AMP; Cysteine; Data; Detection; Diazepam; Dimerization; Dissociation; Disulfides; Dose; Drug Design; Enhancers; Epitopes; Evaluation; Florida; Fluorescence; Funding; G Protein-Coupled Receptor Genes; GTP-Binding Proteins; Generations; Grant; Guanine Nucleotides; Heart Block; Heart Rate; Hydrogen Peroxide; Investigation; Laboratories; Learning; Ligands; Mediating; Modeling; Modification; Molecular; Mutate; Mutation; Neurons; Neuropathy; Numbers; Pain; Peptide Sequence Determination; Pharmaceutical Chemistry; Physiological; Play; Principal Investigator; Purinergic P1 Receptors; Purinoceptor; Reaction; Reading; Reducing Agents; Regulation; Reporter; Research; Research Personnel; Role; Running; Score; Sedation procedure; Seizures; Series; Signal Transduction; Site; Site-Directed Mutagenesis; Slice; Structure; Sulfhydryl Compounds; Synthesis Chemistry; System; Techniques; Therapeutic; Tilia; Time; Tissues; Universities; Vascular Endothelial Growth Factors; Virginia; angiogenesis; authority; base; chronic pain; clinical application; crosslink; disulfide bond; extracellular; improved; insight; interest; macrophage; mathematical model; mutant; novel; prevent; programs; protein activation; prototype; radioligand; receptor; receptor function; research study; response; trafficking

This competitive renewal proposes to continue our investigation of allosteric enhancers (AEs) of the A1 adenosine receptor (A^R). These compounds bind to an allosteric site on the receptor to increase the affinity of adenosine to the orthosteric site on the A^R. We have identified new and improved aminothiophene AEs and we have discovered a new chemical class of more potent and effective aminothiazole AEs. We also have discovered that some disulfides and H2O2 have AE-like effects on the AiAR and the A^R. To learn more about the molecular mechanisms of AE action we investigated receptor mutants and chimera. We have developed a novel system to "score" enhancer activity based on the ability of these compounds to prevent rapid dissociation of an agonist radioligand, 125I-ABA, from the A^R in response to GTPyS. We propose newstudies to achieve three specific aims. Aim 1 is to synthesize new enhancer structures in two chemical classes and to characterize them based on radioligand binding to receptors and guanine nucleotide exchange from G proteins. This aim is guided by conformational molecular field analysis (CoMFA) of existing structures and the use of new synthetic techniques. Aim 2 is to efficiently evaluate enhancer function in intact cells and to evaluate selected compounds for activity in modulating adenosine A1 receptor function in brain slices and stimulation of angiogenesis. Aim 3 will investigate the role and disulfide bond formation in AE and H2O2 action through analysis of thiol alkylating agents, epitope- tagged receptors, site-directed mutagenesis and protein sequencing. We will also investigate the possibility the H2O2 is a physiological regulator of A^R signaling. We hypothesize that cysteines in the extracellular loops of the AiAR participate in disulfide cross linking reactions. AEs are significant therapeutic candidates that may reduce chronic pain or promote site-specific angiogenesis. Moreover, our investigation of the role of thiols/disulfides in enhancer and H2O2 action may reveal important new information about general mechanisms of GPCR G protein coupling and trafficking.

这次竞争性更新建议继续我们对 A1 变构增强剂 (AE) 的研究 腺苷受体（A^R）。这些化合物与受体上的变构位点结合，以增加 腺苷与 A^R 上的正位点的亲和力。我们已经确定了新的和改进的 氨基噻吩 AE，我们发现了一种更有效的新化学类别 氨基噻唑 AE。我们还发现一些二硫化物和H2O2对 AiAR 和 A^R。为了更多地了解 AE 作用的分子机制，我们研究了受体 突变体和嵌合体。我们开发了一种新颖的系统，可以根据增强剂活性的能力“评分” 这些化合物可防止激动剂放射性配体 125I-ABA 从 A^R 中快速解离 对 GTPyS 的反应。我们提出新闻研究来实现三个具体目标。目标1是合成新的 两种化学类别中的增强子结构，并根据放射性配体结合来表征它们 G 蛋白的受体和鸟嘌呤核苷酸交换。这一目标是由构象分子引导的 现有结构的现场分析（CoMFA）和新合成技术的使用。目标 2 是高效 评估完整细胞中的增强子功能并评估所选化合物的调节活性 腺苷 A1 受体在脑切片中的功能和刺激血管生成。目标 3 将调查角色 通过硫醇烷基化剂、表位的分析，了解 AE 和 H2O2 作用中二硫键的形成 标记受体、定点突变和蛋白质测序。我们也会调查可能性 H2O2 是 A^R 信号传导的生理调节剂。我们假设细胞外的半胱氨酸 AiAR 的环参与二硫键交联反应。 AE 是重要的治疗候选者 可以减轻慢性疼痛或促进特定部位的血管生成。此外，我们对角色的调查 增强子中的硫醇/二硫化物和H2O2作用可能揭示有关一般性的重要新信息 GPCR G 蛋白偶联和运输的机制。