STRUCTURAL STUDIES OF ARRESTIN AND RHODOPSIN PHOSPHATASE

拟议研究的长期目标是了解 猝灭相关蛋白质的分子性质 光转导。有三种蛋白质参与了导致 光解视紫红质的失活：视紫红质激酶、arrestin和 视紫红质磷酸酶。首先，视紫红质激酶催化 新漂白的视紫红质的磷酸化启动 光解视紫红质的失活，然后是arrestin和视紫红质 磷酸酶参与了随后的失活步骤。 与激酶相反，只有零碎的信息可用 关于arrestin在光转导中的作用和关于磷酸酶 身份。在许多视网膜变性的病例中，视网膜变性的主要原因 疾病表现为周期性GMT代谢异常或 视紫红质基因的突变；或者，视紫红质基因的变性 当视紫红质水平升高时，感光细胞变得明显 磷酸化/去磷酸化异常。为了定义分子 人类复杂的视网膜变性的基础事件，如 视网膜色素变性，因此光传导的各个步骤必须 在非常详细的水平上被确定。 计划中的实验的目的是(1)阐明 Arrestin的晶体结构。(二)调查情况 当视紫红质的发色团从 磷酸化的膜和视紫红质的脱磷酸化 视紫红质磷酸酶。(3)确定竞争性抑制因子。 视紫红质-arrestin相互作用。(4)为了表征β-arrestin，a Arrestin的同系物，包括其功能和免疫学性质。 (5)明确鉴定视紫红质磷酸酶及其抑制物 它的活动。

The long-term objective of the proposed studies is to understand the molecular properties of the proteins involved in quenching phototransduction. Three proteins are engaged in the process that leads to deactivation of photolyzed rhodopsin: rhodopsin kinase, arrestin and rhodopsin phosphatase. First, rhodopsin kinase catalyzes the phosphorylation of freshly bleached rhodopsin which initiates the deactivation of photolyzed rhodopsin, and then arrestin and rhodopsin phosphatase are involved in the subsequent deactivation steps. In contrast to kinase, only fragmentary information is available about the role arrestin has in phototransduction and about the phosphatase identity. In many cases of retinal degeneration, the primary causes of the disorder appear to be either an abnormality cyclic GMT metabolism or mutations in the rhodopsin gene; alternatively, the degeneration of photoreceptor cells becomes evident when levels of rhodopsin phosphorylation/dephosphorylation are abnormal. To define the molecular events that underlie a complex retinal degeneration in humans, such as retinitis pigmentosa, the various steps of phototransduction therefore must be determined on a very detailed level. The objectives of the planned experiments are (1) to elucidate the crystallographic structure of arrestin. (2) to investigate the status of rhodopsin's chromophore when arrestin is released from the phosphorylated membranes and when rhodopsin is dephosphorylated by rhodopsin phosphatase. (3) to identify competitive inhibitors of the rhodopsin-arrestin interaction. (4) to characterize beta-arrestin, a homolog of arrestin, including its functional and immunological properties. (5) to definitively identify the rhodopsin phosphatase and an inhibitor of its activity.