Multiple Ligand Binding Sites of Rod and Cone Opsins

视杆细胞和视锥细胞视蛋白的多个配体结合位点

• 批准号: 7472444
• 负责人: CLINT L MAKINO
• 金额: $ 37万
• 依托单位: MASSACHUSETTS EYE AND EAR INFIRMARY
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-01-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7472444
• 关键词: 11 cis Retinal; Address; Affect; Agonist; Binding; Binding Sites; Biochemical; Biological Assay; Buffers; Cell physiology; Cells; Cyclic GMP; Darkness; Dependence; Ecology; Energy Transfer; Exposure to; G-Protein-Coupled Receptors; GTP-Binding Proteins; Goals; Kinetics; Ligand Binding; Ligands; Light; Link; Microspectrophotometry; Mutation; Opsin; Pathology; Personal Satisfaction; Pharmacologic Substance; Photons; Photoreceptors; Phototransduction; Physiology; Pigment Epithelium; Pigments; Process; Property; Regulation; Reproducibility; Research Personnel; Rest; Retinal; Retinal Diseases; Retinal Pigments; Retinoids; Rhodopsin; Role; Role playing therapy; Site; Solutions; Spectrum Analysis; Structure; Testing; Vertebrate Photoreceptors; Vision; Vision research; Visual; absorption; analog; chromophore; cyclic-nucleotide gated ion channels; improved; programs; protein expression; receptor; research study; response; retinal rods; visual cycle; visual process; visual processing

DESCRIPTION (provided by applicant): Many G protein coupled receptors, visual opsins included, weakly activate G protein in the absence of ligand. Opsins are unique in their binding of an inverse agonist at rest, 11-cis retinal, which is transformed into an agonist by light. The interactions between rod opsin and its 11-cis retinal chromophore have been well studied for their roles in pigment stability, activation and spectral tuning. Recently, it has been proposed that opsin possesses two additional retinoid binding sites that enable retinoids to approach and exit the pigment-forming chromophore binding pocket. In this study, we will provide further evidence for multiple ligand binding sites on opsin. More importantly, we hypothesize that opsins bind other ligands besides retinoids and that the functions of the binding sites extend beyond visual cycle. Using single cell physiology, single cell microspectrophotometry and spectroscopy on rhodopsin in solution, we propose experiments to test three functions. The light-regulated channel is susceptible to potent inhibition by retinoids. One function of the alternate binding sites may be to allow rhodopsin and opsin to serve as buffers that protect the cGMP-gated channel from retinoid inhibition after exposure to bright light, when vast amounts of retinoids are exchanged with the pigment epithelium. A second function of retinoid binding to alternate sites is to modulate the catalytic activity of rhodopsin and opsin. This function may play roles in conferring reproducibility to the single photon response, in setting the absolute sensitivity and response kinetics of the receptor. A third function may be to enable compounds to absorb light and transfer the energy to rhodopsin, causing the latter to photoisomerize and give rise to an electrical response as if by direct absorption. Thus spectral sensitivity may be altered without a change in opsin protein expression. Each of these functions may depend upon opsin type. A strong link has been established between phototransduction cascade activity and retinal disease raising the possibility that mutations affecting the alternate ligand binding sites or that affect ligand processing may be responsible for or contribute to certain retinal pathologies. Then pharmaceuticals that target the alternate site may be used for the treatment of a retinal disease or for the enhancement of vision. Therefore, this proposal will be highly significant to our understanding of the structure and function of visual opsins and opens new directions for vision research.

描述（由申请人提供）：许多G蛋白偶联受体，包括视觉视蛋白，在没有配体的情况下微弱地激活G蛋白。视蛋白的独特之处在于其在静止时与反向激动剂 11-顺式视网膜结合，在光作用下将其转化为激动剂。杆状视蛋白与其 11-顺式视网膜发色团之间的相互作用因其在色素稳定性、激活和光谱调谐中的作用而得到了深入研究。最近，有人提出视蛋白拥有两个额外的类视黄醇结合位点，使类视黄醇能够接近和离开形成色素的发色团结合袋。在这项研究中，我们将为视蛋白上的多个配体结合位点提供进一步的证据。更重要的是，我们假设视蛋白除了类视黄醇外还结合其他配体，并且结合位点的功能超出了视觉周期。利用单细胞生理学、单细胞显微分光光度法和溶液中视紫红质的光谱学，我们提出了测试三种功能的实验。光调节通道容易受到类维生素A的有效抑制。替代结合位点的一项功能可能是允许视紫红质和视蛋白充当缓冲剂，在暴露于强光后，当大量类维生素A与色素上皮交换时，保护cGMP门控通道免受类维生素A抑制。类维生素A与替代位点结合的第二个功能是调节视紫红质和视蛋白的催化活性。该功能可能在赋予单光子响应的再现性、设定受体的绝对灵敏度和响应动力学方面发挥作用。第三个功能可能是使化合物能够吸收光并将能量转移到视紫红质，导致后者光致异构化并产生电响应，就像直接吸收一样。因此，可以在不改变视蛋白表达的情况下改变光谱灵敏度。这些功能中的每一个都可能取决于视蛋白类型。光转导级联活性和视网膜疾病之间已经建立了密切的联系，这提高了影响替代配体结合位点或影响配体加工的突变可能导致或促成某些视网膜病理的可能性。然后，针对替代部位的药物可用于治疗视网膜疾病或增强视力。因此，该提议对于我们理解视觉视蛋白的结构和功能具有重要意义，并为视觉研究开辟新的方向。