权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Structural analysis and functional expression of vasopressin water channel

加压素水通道的结构分析及功能表达

基本信息

批准号：
07671243
负责人：
FUSHIMI Kiyohide
金额：
$ 1.47万
依托单位：
Tokyo Medical and Dental University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1995
资助国家：
日本
起止时间：
1995 至 1996
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-07671243/
关键词：
AQP2 aquaporin-2 water channel vasopressin collecting duct urine concentration trafficking exocytosis

项目摘要

Methods of molecular biotechnology has been applied to study molecular structure of aquaporin-2 vasopressin water channel. Parts of potentially critical structure of aquaporin-2 were replaced with corresponding parts of other MIP family channels, including GlpF,a glycerol facilitator of E.Coli, and MIP,a non-specific channel of lens. Chimeras were expressed in Xenopus oocytes and mammalian epithelial cells, and structure-function relations of parts of AQP2 were analyzed. It was shown that the aqueous pore of AQP2 was assembled with aligned six transmembrane segments in alpha-helical structure. A constriction that passes only water molecules, thus accounting for selective water permeability, was created with the third and fourth hydrophilic loops in ectoplasmic and endoplasmic side respectively.In addition, functional expression studies have elucidated that the structure of the endoplasmic domain of AQP2 carboxyl terminus is critically important for plasma membrane expression of AQP2. Especially, it was shown that the carboxyl terminus plays crucial roles in vasopressin-regulated translocation of AQP2, a key feature of vasopressin antidiuresis in the collecting duct cells as revealed in previous studies. The serine 256 in AQP2 carboxyl terminus is phosphorylated by protein kinase A and a mutation in the serine 256 diminished vasopressin-regulated exocytosis of AQP2. Thus, it was proved that phosphorylation of the serine 256 is required for vasopression-regulated trafficking of AQP2. This finding is interesting in that vesicle trafficking system is not only regulated by vesicle-associated proteins but also regulated through modifications on AQP2, a major cargo of vesicles.

应用分子生物技术方法研究了aquaporin-2加压素水通道的分子结构。 aquaporin-2 的潜在关键结构部分被其他 MIP 家族通道的相应部分取代，包括 GlpF（大肠杆菌的甘油促进剂）和 MIP（晶状体非特异性通道）。嵌合体在爪蟾卵母细胞和哺乳动物上皮细胞中表达，并分析了 AQP2 部分的结构-功能关系。结果表明，AQP2 的水孔由排列成 α 螺旋结构的六个跨膜片段组装而成。分别在外质侧和内质侧的第三和第四亲水环产生了仅通过水分子的收缩，从而实现了选择性水渗透。此外，功能表达研究表明，AQP2羧基末端的内质结构域的结构对于AQP2的质膜表达至关重要。特别是，研究表明，羧基末端在加压素调节的 AQP2 易位中起着至关重要的作用，这是先前研究揭示的集合管细胞中加压素抗利尿作用的关键特征。 AQP2 羧基末端的丝氨酸 256 被蛋白激酶 A 磷酸化，丝氨酸 256 的突变减少了 AQP2 加压素调节的胞吐作用。因此，证明丝氨酸256的磷酸化是血管加压调节的AQP2运输所必需的。这一发现很有趣，因为囊泡运输系统不仅受到囊泡相关蛋白的调节，而且还通过对囊泡的主要货物 AQP2 的修饰进行调节。