Aquaporin 0 calmodulin interaction and the effect of Aquaporin 0 phosphorylation
Aquaporin 0 calmodulin interaction and the effect of Aquaporin 0 phosphorylation
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DOI：
10.1021/bi701980t
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发表时间：
2008-01-08
期刊：
影响因子：
2.9
通讯作者：
Schey, K. L.
中科院分区：
文献类型：
--
作者：
Rose, K. M. Lindsey;Wang, Z.;Schey, K. L.
水孔蛋白0 （Aquaporin 0, AQP0）又称晶状体的主要内在蛋白，是晶状体中含量最多的膜蛋白，它经历了一系列c端导向的翻译后修饰。含有主要磷酸化位点的c端区域是一个假定的钙调素结合位点，钙调素已被证明调节AQP0的水渗透性。本研究的目的是阐明AQP0磷酸化在钙调蛋白结合中的作用。在S231和S235上分别合成了丝氨酸磷酸化和不磷酸化的AQP0 c端肽，并利用荧光结合试验评估了这些肽与丹酚标记的钙调素的结合能力以及相互作用对钙的依赖性。发现AQP0 c端磷酸化肽对钙调素的亲和力比未磷酸化肽低20-50倍。化学交联研究揭示了AQP0-钙调蛋白相互作用的特定位点被AQP0磷酸化显著降低。这些数据提示AQP0 c端磷酸化在体内影响钙调蛋白结合，并对AQP0功能有调控作用。
Aquaporin 0 (AQP0), also known as major intrinsic protein of lens, is the most abundant membrane protein in the lens and it undergoes a host of C-terminally directed posttranslational modifications. The C-terminal region containing the major phosphorylation sites is a putative calmodulin-binding site, and calmodulin has been shown to regulate AQP0 water permeability. The purpose of the present study was to elucidate the role of AQP0 phosphorylation on calmodulin binding. AQP0 C-terminal peptides were synthesized with and without serine phosphorylation on S231 and S235, and the ability of these peptides to bind dansyl-labeled calmodulin and the calcium dependence of the interaction was assessed using a fluorescence binding assay. The AQP0 C-terminal phosphorylated peptides were found to have 20-50-fold lower affinities for calmodulin than the unphosphorylated peptide. Chemical cross-linking studies revealed specific sites of AQP0-calmodulin interaction that are significantly reduced by AQP0 phosphorylation. These data suggest that AQP0 C-terminal phosphorylation affects calmodulin binding in vivo and has a role in regulation of AQP0 function.