SYNTHETIC COMPOUNDS TO PROBE MOLECULAR EVENTS IN BIOLOGY

用于探测生物学分子事件的合成化合物

• 批准号: 3734434
• 负责人: VALERIA BALOGH-NAIR
• 金额: --
• 依托单位: CITY COLLEGE OF NEW YORK
• 依托单位国家: 美国
• 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/3734434
• 关键词: G protein; Raman spectrometry; chemical binding; chemical structure function; chemical synthesis; chromophore; high performance liquid chromatography; photochemistry; protein structure; rhodopsin; spectrometry; stereochemistry; visual photoreceptor

We propose an organochemical approach to determine molecular recognition elements that the visual photoreceptors employ to trigger the cascade of biochemical events, which culminate in neural responses. We are specifically interested in: 1) the nature of molecular events at the retinal chromophore's binding site in rhodopsins that enables the ultrafast photoisomerization and wavelength regulation, 2) the mechanisms of conformational changes of the rhodopsin, initiated by retinal isomerization, that in turn create a binding domain in the tertiary structure of rhodopsin, structurally optimized for photoactivated rhodopsin - G protein (transducin) interactions. Concerted efforts of several disciplines have been successful in elucidating many aspects of this information processing system but the tertiary structures of rhodopsin and its photoactivated form are not known. Knowledge of these tertiary structures are however, a prerequisite for understanding the mechanism of the sequence of events in a structural perspective, at the molecular level, 3) the structural make-up of the G protein binding site that requires guanine nucleotides and hitherto was difficult to investigate due to the lack of appropriately labeled nucleotide analogs. We propose to achieve our goals by incorporation of synthetic probes into rhodopsins and G proteins followed by studies of the properties of functional rhodopsin and G protein analogs generated. In the case of rhodopsin, novel type of spacer-armed retinals will be employed, and in the case of G protein, isotopically labeled and site specifically modified GDPs will be used as probes of the protein's structure. The structural information obtained from employing the probes described in this project will open up a whole new area of exploratory studies. For instance, development of domain-specific rhodopsin-immunoconjugates and site-specific rhodopsin-peptide conjugates will be possible. It will also pave the way to site-specific conjugation of rhodopsins to effect their crystallization and to effect their site-specific cleavage. In the case of the G protein, Raman spectroscopic investigation employing isotopically labeled GDP analogs will yield insight into the solution structure of the interaction domain(s) of the G with the protein. The organochemical approach in this project will thus have a major impact in the area of sensory transduction, where molecular level investigations can make unique contributions towards the elucidation of the cellular events in precise structural terms, by identifying the molecular domains and chemical mechanisms involved in the activation of excitable membranes.

我们提出了一种有机化学方法来确定分子识别 视觉感光器用来触发级联的元素 生化事件，最终导致神经反应。 我们是 特别感兴趣的是：1）分子事件的性质 视紫红质中视网膜发色团的结合位点，使 超快光异构化和波长调节，2）机制 由视网膜引发的视紫红质构象变化 异构化，进而在三级结构中创建结合域 视紫红质的结构，针对光激活进行了结构优化 视紫红质 - G 蛋白（转导蛋白）相互作用。 齐心协力 一些学科已经成功地阐明了 这个信息处理系统而是三级结构 视紫红质及其光激活形式尚不清楚。 这些知识 然而，三级结构是理解 从结构角度看事件发生顺序的机制 分子水平，3）G蛋白结合位点的结构组成 这需要鸟嘌呤核苷酸，而迄今为止很难做到 由于缺乏适当标记的核苷酸类似物而进行调查。 我们建议通过将合成探针纳入其中来实现我们的目标 视紫红质和 G 蛋白的特性研究 生成功能性视紫红质和G蛋白类似物。 如果是 将采用视紫红质，新型间隔臂视网膜，并在 以 G 蛋白为例，经过同位素标记和位点特异性 修改后的 GDP 将用作蛋白质结构的探针。 这 通过使用中描述的探针获得的结构信息 该项目将开辟一个全新的探索性研究领域。 为了 例如，开发域特异性视紫红质免疫缀合物和 位点特异性视紫红质-肽缀合物将成为可能。 它会 也为视紫红质的位点特异性结合铺平道路 它们的结晶并实现其位点特异性裂解。 在 G 蛋白的情况，拉曼光谱研究采用 同位素标记的 GDP 类似物将有助于深入了解解决方案 G 与蛋白质的相互作用结构域的结构。 因此，该项目中的有机化学方法将产生重大影响 在感觉转导领域，分子水平的研究 可以为阐明细胞做出独特的贡献 通过识别分子域，以精确的结构术语描述事件 以及参与兴奋性激活的化学机制 膜。