CYSTIC FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR CHLORIDE CHANNEL

囊性纤维化跨膜电导调节剂氯离子通道

• 批准号: 6307561
• 负责人: DAVID C GADSBY
• 金额: $ 0.82万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-12-01 至 2000-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6307561
• 关键词: biomedical resource; human tissue; nervous system; spectrometry

The disease cystic fibrosis results from reduced epithelial Cl-permeability due to mutations in the gene encoding the cystic fibrosis traiismembrane conductances regulator (CFTR) Cl-channel. CFTR channels, like all other members of the family of ATP-binding cassette (ABC) transporters, incorporates two nucleotide binding domains (NBDs) but also includes a unique regulatory R domain containing more than 10 consensus sites for phosphorylation by cAMP-dependent protein kinase (PKA) and protein kinase C (PKC). In cells with ' normal CFTR channels, receptor-mediated activation of PKA causes phosphorylation of several R-domain serines, permitting channel opening and closing via cycles involving ATP hydrolysis. We have recently obtained strong evidence that ATP hydrolysis energizes the conforinational change that opens the channel gate, and that the degree of phosphorylation of a channel is one of the determinants of how long the gate stays open. Our working hypothesis is that phosphorylation of particular serines controls, independently, the function of the two NBDs. In a strongly phosphorylated channel with both NBDs functional, then hydrolysis of ATP at one NBD opens the channel, whereupon a second ATP can bind at the other NBD and in so ding can stabilize the open conformation. Hydrolysis of that second ATP then abolishes the stabilization, prompting channel closure. We have also hypothesized that distinct cellular phosphatases differentially dephosphorylate the various phosphoserines. Hence, in the cell, activation or inhibition of specific phosphatases could contribute to the complex mechanisms that regulate channel gating. The aim of this project is to learn which serines are phosphorylated under which experimental condition, with the goal of eventually discerning the exact role of each phosphoserine in orchestrating the function of the individual channel domains. The approach is to correlate biochemical information on phosphorylation with precise assays of function at the single channel level.

疾病囊性纤维化是由上皮减少引起的 由于编码囊性的基因中的突变而引起的Cl-渗透性 纤维化Traiismmbrane电导调节剂（CFTR）CL通道。 CFTR频道，与ATP结合家庭的所有其他成员一样 盒式转运蛋白包含两个核苷酸结合 域（NBD），但还包括一个独特的调节域 包含超过10个共识位点，用于磷酸化 cAMP依赖性蛋白激酶（PKA）和蛋白激酶C（PKC）。 在 具有正常CFTR通道的细胞，受体介导的PKA激活 引起几个R域丝氨酸的磷酸化，允许通道 通过涉及ATP水解的周期开放和关闭。 我们有 最近获得了有力的证据，表明ATP水解能量能量 授权的变革开放了渠道大门， 通道的磷酸化程度是一种决定因素之一 大门留下多长时间。 我们工作的假设是 特定丝氨酸控制的磷酸化独立于 两个NBD的功能。 在强烈磷酸化的通道中 NBD的两个功能，然后在一个NBD下对ATP的水解打开 频道，因此第二个ATP可以在其他NBD上绑定，因此 丁可以稳定开放构象。 第二个水解 然后，ATP废除了稳定，促使通道闭合。 我们 还假设不同的细胞磷酸酶 差异化各种磷酸盐。 因此，在 细胞，激活或抑制特定磷酸酶可以 有助于调节通道门控的复杂机制。 该项目的目的是学习哪些丝氨酸是磷酸化的 在哪个实验条件下，最终的目标 辨别每种磷serine在策划的确切作用 单个通道域的功能。 方法是 与精确的磷酸化的生化信息相关 在单个通道级别的功能测定。