SYNAPTIC VESICLE CATECHOLAMINE/AEROTINA

突触小泡儿茶酚胺/Aerotina

• 批准号: 3401368
• 负责人: JOSEPH A NEAR
• 金额: $ 8.78万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-04-01 至 1992-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3401368
• 关键词: adenosine triphosphate; affinity chromatography; affinity labeling; biomarker; brain cell; catecholamine inhibitor; catecholamines; central nervous system; chemical association; chemical binding; chromaffin cells; cow; crosslink; electrochemistry; electrophoresis; glycoproteins; isoelectric point; molecular weight; neurochemistry; radioassay; reserpine; serotonin; synaptic vesicles; tetrabenazine; tritium

The accumulation of catecholamines and serotonin in synaptic vesicles of the CNS is mediated by a transport system (porter) located in the vesicle membrane. This system is the primary target for several pharmacological agents that have been used extensively in research and are of considerable clinical interest. The long- term objectives of the proposed research include identification, reconstitution and characterization of the molecular entities directly involved in transport. Of particular interest are the subunit structure of the porter, and localization and orientation of binding sites for inhibitors such as reserpine and tetrabenazine. These two agents appear to represent distinct classes of inhibitors that may interact with different subunits of the porter complex in order to achieve these goals photoaffinity analogs of tetrabenazine, reserpine and substrates will be used to identify relevant components of the complex ln synaptic vesicles from beef brain, and labeled components will be characterized with respect to molecular weight isoelectric point, and glycoprotein content by lectin affinity chromatography. The effect of membrane energization on photoaffinity labeling will also be assessed. The respective binding components will be solubilized and purified by conventional biochemical methods, including affinity chromatography using inhibitor binding or photoaffinity labeling to follow activity. The ability of purified components to bind substrates and inhibitors will be compared, and the pH dependence of such binding assessed. The type of inhibition exerted by various agents on dopamine transport and reserpine and dihydrotetrabenazine binding to purified components will be examined. In addition to a better understanding of the structure and organization of the porter, the proposed work should yield information useful in the design of new catecholamine depleting therapeutic agents with potential utility in clinical situations that require a reduction of catecholamine available for release to the synapse.

突触内儿茶酚胺和5-羟色胺的积累 中枢神经系统的囊泡由转运系统（搬运工）介导 位于囊泡膜中。 这个系统是首要目标 对于几种已经广泛使用的药理学试剂， 在研究中，并具有相当大的临床意义。 很长的- 拟议研究的长期目标包括识别， 分子实体的重构和表征 直接参与运输。 特别感兴趣的是 转运蛋白的亚基结构、定位和定向 利血平等抑制剂的结合位点 丁苯那嗪 这两个代理人似乎代表了不同的 可能与不同亚基相互作用的抑制剂类别 为了实现这些目标， 丁苯那嗪、利血平和底物的类似物将用于 识别突触囊泡中复合物的相关成分 来自牛脑，标记的成分将被表征为 相对于分子量等电点，和糖蛋白 含量通过凝集素亲和层析法测定。 膜的作用 还将评估双链体光亲和标记。 的 通过以下方法溶解和纯化各个结合组分 常规生物化学方法，包括亲和色谱法 使用抑制剂结合或光亲和标记， 活动 纯化组分结合底物的能力 和抑制剂将进行比较，并且这种pH依赖性 约束力评估 各种药剂产生的抑制类型 多巴胺转运和利血平以及二氢丁苯那嗪 将检查与纯化组分的结合。 除了 更好地了解联合国的结构和组织， 波特，拟议的工作应该产生有用的信息， 设计新的消耗儿茶酚胺的治疗剂， 在需要减少的临床情况下的潜在效用 释放到突触中。