MOLECULAR MECHANISMS IN VISUAL TRANSDUCTION

视觉传导的分子机制

• 批准号: 3258023
• 负责人: ROBERT R RANDO
• 金额: $ 15.21万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-12-01 至 1991-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3258023
• 关键词: Schiff bases; acidity /alkalinity; aminoacid; bioenergetics; biological signal transduction; carboxyl group; chemical structure function; chromophore; cis trans isomerization; conformation; crosslink; drug receptors; electron microscopy; guanosine triphosphate; lysine; methylation; molecular site; photochemistry; photolysis; protonation; radioassay; rhodopsin; spectrometry; transducin; vision; visual pigments

Our long-term objectives are to understand the molecular mechanisms of visual transduction and adaptation. This proposal is focused on the molecular mechanism of rhodopsin action. After rhodopsin absorbs a photon it proceeds through a series of spectroscopically defined conformation states. One or more of these conformational states (R*) catalyzes the exchange of GTP for GDP in a disk associated G-protein resulting in the activation of the latter. Similar, if not identical , informational transducing steps occur as a consequences of the interaction of many hormones and drugs with their receptors. The proposal is largely concerned with defining the molecular changes in rhodopsin which enable activated rhodopsin R* to be reached. A major interest here is to establish a structural link between R* and the spectroscopically defined intermediates. This work will be of relevance both to our understanding of vision and the nature of drug-receptor interactions. The fact that Schiff base of rhodopsin can be protonated is of immense importance in the functioning of this protein. The protonated Schiff base of the chromophore, interacting with negatively charge amino acids at the active-site, is thought to be of primary importance in wavelength regulation and energization of rhodopsin. The ultimate deprotonation of the Schiff base is assumed also to be important in the formation of metarhodopsin II, the presumed spectroscopic signature of R*. Along these lines, the active-site lysine of opsin must also be deprotonated to form a Schiff base with 11-cis-retinal. The experiments described here are designed to prove the role of charge and charge movement in the functioning of rhodopsin and, to a much lesser extent, bacteriorhodopsin. The approaches used here are bio-organic chemical and biochemical in nature and will involve modification of the protein along with retinal analog studies. By introducing chemical probes, such as a methyl group, at the active-site lysine we will determine if a full positive charge on this lysine is required for photochemical energy storage, wavelength regulation and the formation of R*. Furthermore the pK of the active-site lysine of rhodopsin will be determined. The amino acid counterions which critically interact with the chromophore will be identified by structural studies designed to use the active-site lysine of opsin to direct a pseudo cross-linking reagent to these counterions. Finally, novel retinal analogs which form pigments with opsin will be used to probe the mechanism of energy storage.

我们的长期目标是了解分子 视觉转导和适应的机制。 这个建议 专注于视紫红质作用的分子机制。 后 Rhodopsin吸收光子通过一系列的光子进行 光谱定义的构象状态。 一个或多个 这些构象状态（R*）催化GTP的交换 用于磁盘中的GDP相关的G蛋白，导致激活 后者。 相似，即使不是完全相同的信息传递 步骤是由于许多人相互作用而发生的 激素和药物及其受体。 该提议主要是 与定义视紫红质的分子变化有关，该变化 启用激活的视紫红质R*。 主要兴趣 这是建立r*和 光谱定义的中间体。 这项工作将是 与我们对视觉的理解和本质有关 药物受体相互作用。 Rhodopsin的Schiff基础可以质子化的事实是 该蛋白质的功能极为重要。 这 发色团的质子化的席夫碱，与 据认为，在活性位点负电荷氨基酸是 波长调节和能量的主要重要性 Rhodopsin。 Schiff基地的最终去质子化是 假定在形成metarhopopsin中也很重要 II，R*的假定光谱特征。 沿着这些线 OPSIN的活性位点赖氨酸也必须被质子质子化以形成 带有11盘视网膜的Schiff基地。 这里描述的实验 旨在证明充电和电荷运动的作用 视紫红质的功能，并且在较小程度上的功能 细菌淡淡蛋白质。 这里使用的方法是生物有机 化学和生化本质上，将涉及修改 蛋白质以及视网膜模拟研究。 通过介绍 化学探针，例如甲基，在活性位点赖氨酸处 我们将确定该赖氨酸的全部正电荷是 需要光化学能量存储，波长调节 和r*的形成。 此外，活动点的PK 将确定Rhodopsin的赖氨酸。 氨基酸 与发色团严重相互作用的对抗将是 通过旨在使用活性站点的结构研究确定 Opsin的赖氨酸将伪交联试剂引导到这些试剂 反击。 最后，形成颜料的新型视网膜类似物 使用OPSIN将用于探测能量储能的机理。