MOLECULAR MECHANISMS IN VISUAL TRANSDUCTION

视觉传导的分子机制

• 批准号: 3258025
• 负责人: ROBERT R RANDO
• 金额: $ 15.05万
• 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/3258025
• 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 视觉传导和适应机制。 这项建议 主要研究视紫红质作用的分子机制。 后 视紫红质吸收一个光子， 光谱学定义的构象状态。 中的一个或多 这些构象状态（R*）催化GTP的交换 对于椎间盘中与G蛋白相关的GDP， 后者。 类似的，如果不是完全相同的，信息转导 步骤的发生是许多因素相互作用的结果， 激素和药物及其受体。 该提案主要是 研究视紫红质的分子变化， 使活性视紫红质R* 达到。 主要兴趣 这里是建立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 药物-受体相互作用 视紫红质的席夫碱可以质子化的事实是 对这种蛋白质的功能有着巨大的影响。 的 发色团的质子化席夫碱，与 在活性位点的负电荷氨基酸，被认为是 在波长调节和波长转换中具有首要重要性 视紫红质 席夫碱的最终去质子化是 也被认为是重要的形成变视紫红质 II，R* 的假定光谱特征。 沿着这些路线， 视蛋白活性位点赖氨酸也必须去质子化以形成 带有11-顺式-视黄醛的席夫碱。 这里描述的实验 旨在证明电荷和电荷运动在 视紫红质的功能，在很小的程度上， 细菌视紫红质 这里使用的方法是生物有机的 化学和生物化学性质，并将涉及修改 沿着视网膜类似物的研究。 通过引入 化学探针，如甲基，在活性位点赖氨酸 我们将确定这个赖氨酸上的全部正电荷是否 光化学能量存储、波长调节所需的 以及R* 的形成。 此外，活性位点的pK 将测定视紫红质的赖氨酸。 的氨基酸 与发色团临界相互作用的抗衡离子将是 通过设计使用活性位点的结构研究确定 将假交联剂引导至这些 反离子 最后，形成色素的新型视网膜类似物 与视蛋白的相互作用将被用来探索能量储存的机制。