CRYSTALLOGRAPHIC STUDIES OF THE AQP0-CAM COMPLEX

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 我们研究了水通道蛋白0(AQP0)，这是一种来自眼睛晶状体的跨膜水通道，当突变时会导致晶状体严重损伤、白内障和失明。AQP0通过与钙调蛋白相互作用，对[Ca~(2+)]的变化作出反应，从而调节眼晶状体内的水分渗透性。钙调素(CaM)是一种普遍存在的胞浆蛋白，常与钙离子调节偶联。我们认为，钙/钙调素与AQP0的相互作用导致水通道内的主要构象变化，从而影响水的通透性。这项建议的目的是利用X射线结晶学来确定AQP0-CaM络合物的高分辨率结构。我们可以疲倦地生长同时含有AQP0和CaM的大晶体，但由于衍射率低，高分辨率结构信息的获得一直受到阻碍。成功地实现我们的目标将在水通道调控领域产生广泛的影响，同时也将对钙/钙调素信号系统和膜结构生物学的进一步发展产生更广泛的影响。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. We study aquaporin 0 (AQP0), a transmembrane water channel from the eye lens, causing severe lesions in the lens, cataracts and blindness when mutated. AQP0 regulates water permeability within the eye lens by responding to changes in [Ca2+] through an interaction with calmodulin. Calmodulin (CaM) is a ubiquitous cytosolic protein commonly coupled with Ca2+ regulation. We suggest the Ca2+/CaM interaction with AQP0 lead to major conformational changes within the water channels that enfluence water permeability rates. The aims of this proposal are to use X-ray crystallography to determine the high-resolution structure of the AQP0-CaM complex. We can fatihfully grow large crystals containing both AQP0 and CaM, however obtaining high-resolution structural information has been stymied by poor diffraction. Success in achieving our aim will have wide implications in the fields of water channel regulation, as well as broader implications to the Ca2+/CaM signaling system and to furthering membrane structural biology in general.