Functional analysis of the Na+,K+/H+ antiporter PeNHX3 from the tree halophyte Populus euphratica in yeast by model-guided mutagenesis.
Functional analysis of the Na+,K+/H+ antiporter PeNHX3 from the tree halophyte Populus euphratica in yeast by model-guided mutagenesis.
复制标题
模型引导诱变对盐生植物胡杨 Na , K /H 逆向转运蛋白 PeNHX3 的功能分析
DOI：
10.1371/journal.pone.0104147
复制
发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Qiu QS
中科院分区：
文献类型：
--
作者：
Wang L;Feng X;Zhao H;Wang L;An L;Qiu QS
Na+、K+/H+逆向转运蛋白是H+偶联的共转运蛋白，对细胞内环境的稳定至关重要。胡杨是一种著名的盐生树种，含有6个Na+/H+逆向转运蛋白基因(PeNHX1-6)，已被证明在耐盐性中起作用。然而，控制其离子传输的催化机制在很大程度上仍不清楚。以大肠杆菌Na+/H+逆向转运蛋白(ECNhaA)的晶体结构为模板，构建了由胡杨出发的PeNHX3的三维结构。PeNHX3模型显示了典型的TM4-TM11组装，这对离子结合和转运至关重要。PeNHX3的结构遵循“正-内”规律，表现出典型的转运蛋白的物理化学性质。在PeNHX3的TM4-TM11装配区发现了4个保守残基，包括Tyr149、Asn187、Asp188和Arg356。突变分析表明，这些保留的残基是PeNHX3功能所必需的：Asn187和Asp188(形成一个ND基序)控制离子的结合和转运，Tyr149和Arg356补偿TM4-TM11组装中的螺旋偶极。在酵母生长实验中，PeNHX3介导了Na+、K+和Li+的转运。结构域切换分析表明，TM11在Li+转运中起着至关重要的作用。讨论了PeNHX3在离子结合和移位方面的新特点。
Na+,K+/H+ antiporters are H+-coupled cotransporters that are crucial for cellular homeostasis. Populus euphratica, a well-known tree halophyte, contains six Na+/H+ antiporter genes (PeNHX1-6) that have been shown to function in salt tolerance. However, the catalytic mechanisms governing their ion transport remain largely unknown. Using the crystal structure of the Na+/H+ antiporter from the Escherichia coli (EcNhaA) as a template, we built the three-dimensional structure of PeNHX3 from P. euphratica. The PeNHX3 model displays the typical TM4-TM11 assembly that is critical for ion binding and translocation. The PeNHX3 structure follows the ‘positive-inside’ rule and exhibits a typical physicochemical property of the transporter proteins. Four conserved residues, including Tyr149, Asn187, Asp188, and Arg356, are indentified in the TM4-TM11 assembly region of PeNHX3. Mutagenesis analysis showed that these reserved residues were essential for the function of PeNHX3: Asn187 and Asp188 (forming a ND motif) controlled ion binding and translocation, and Tyr149 and Arg356 compensated helix dipoles in the TM4-TM11 assembly. PeNHX3 mediated Na+, K+ and Li+ transport in a yeast growth assay. Domain-switch analysis shows that TM11 is crucial to Li+ transport. The novel features of PeNHX3 in ion binding and translocation are discussed.
登录
查看更多内容
影响因子：
3.3
作者：
Bowers, K;Levi, BP;Stevens, TH
通讯作者：
Stevens, TH
影响因子：
3.5
作者：
INOUE, H;NOUMI, T;KANAZAWA, H
通讯作者：
KANAZAWA, H
影响因子：
11.6
作者：
Bassil, Elias;Ohto, Masa-aki;Blumwald, Eduardo
通讯作者：
Blumwald, Eduardo
影响因子：
64.8
作者：
Fukada-Tanaka, S;Inagaki, Y;Iida, S
通讯作者：
Iida, S
影响因子：
7.2
作者：
Leidi, Eduardo O.;Barragan, Veronica;Pardo, Jose M.
通讯作者：
Pardo, Jose M.