Ion transport and ligand binding by the Na-K-Cl cotransporter, structure-function studies
Ion transport and ligand binding by the Na-K-Cl cotransporter, structure-function studies
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DOI：
10.1016/s1095-6433(01)00420-2
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发表时间：
2001-10-01
影响因子：
2.3
通讯作者：
Forbush, B
中科院分区：
文献类型：
--
作者：
Isenring, P;Forbush, B
阳离子-Cl协同转运体（CCCs）介导Na和/或K与Cl跨质膜的偶联运动。迄今为止，已鉴定出8种CCC：2种Na-K-Cl共转运蛋白（NKCC）、4种K-Cl共转运蛋白（KCCs）、1种Na-Cl共转运蛋白（NCC）和1种CCC相互作用蛋白（CIP）。所有的NKCC和KCC都被袢利尿剂抑制;汞和其他修饰剂也已知会阻断NKCC介导的转运。在这项工作中，我们利用突变的方法来研究不同底物和NKCC之间的相互作用。我们依赖于功能不同的载体（鲨鱼NKCC 1和人类NKCC 1）之间的交换域的策略，以确定参与与离子，环利尿剂和汞的相互作用的残基或残基组。我们的研究结果表明，N-和C-末端在确定离子转运和布美他尼结合的物种差异中没有作用。另一方面，汞和NKCCs之间的相互作用被发现部分涉及的C-末端通过残基，含有可用的巯基。在跨膜片段中，第2、第4和第7预测的α-螺旋中的变体残基被示出编码鲨鱼和人类协同转运蛋白之间的离子转运差异。对于袢利尿剂结合，似乎涉及整个中央结构域的几个区域。有趣的是，这些区域与参与阳离子或阴离子运输以及汞结合的区域不同。(C)2001 Elsevier Science Inc. All rights reserved.
The cation-CI cotransporters (CCCs) mediate the coupled movement of Na and/or K to that of Cl across the plasmalemma of animal cells. Eight CCCs have been identified to date: two Na-K-Cl cotransporters (NKCC), four K-Cl cotransporters (KCCs), one Na-Cl cotransporter (NCC) and one CCC interacting protein (CIP). All of the NKCCs and KCCs are inhibited by loop diuretics; mercury and other modifying agents are also known to block NKCC-mediated transport. In this work, we have utilized a mutational approach to study the interaction between different substrates and the NKCCs. We relied on the strategy of exchanging domains between functionally distinct carriers (the shark NKCC1 and the human NKCC1) to identify residues or group of residues that are involved in the interaction with ions, loop diuretics and Hg. Our results show that the N- and C-termini have no role in determining the species differences in ion transport and bumetanide binding. On the other hand, the interaction between Hg and the NKCCs is found to partially involve the C-terminus through residues that contain available sulfhydryl groups. Within the transmembrane segments, variant residues in the 2nd, 4th and 7th predicted alpha -helices are shown to encode the differences in ion transport between the shark and the human cotransporters. For loop diuretic binding, several regions throughout the central domain appear to be involved. Interestingly, these regions are not the same as those involved in cation or anion transport, and in Hg binding. (C) 2001 Elsevier Science Inc. All rights reserved.