Molecular determinants of permeation through the cation channel TRPV4
Molecular determinants of permeation through the cation channel TRPV4
复制标题
DOI：
10.1074/jbc.m204828200
复制
发表时间：
2002-09-13
影响因子：
4.8
通讯作者：
Nilius, B
中科院分区：
文献类型：
--
作者：
Voets, T;Prenen, J;Nilius, B
我们研究了离子通过 TRPV4 通道（VRL-2、TRP12、VR-OAC 和 OTRPC4）渗透的分子决定因素。 TRPV4 的特点是向内和向外整流、钌红的电压依赖性阻断、对二价与单价阳离子的中等选择性以及艾森曼 IV 渗透性序列。我们确定了两个天冬氨酸残基 Asp(672) 和 AsP682，它们是 TRPV4 孔 Ca2+ 敏感性的重要决定因素。天冬氨酸与丙氨酸的中和导致二价阳离子的相对渗透性和向外精馏程度的适度降低。同时中和两种天冬氨酸会导致 Ca2+ 渗透性和通道整流更强烈地降低，并另外改变单价阳离子对艾森曼序列 II 或 I 的渗透性顺序。此外，中和 Asp(682) 但不中和 AsP672 会强烈降低通道对钌红的亲和力。 Met(680) 的突变位于假定的选择性过滤器的中心，会强烈降低全细胞电流幅度并损害 Ca2+ 渗透。相比之下，中和假定孔区域中唯一带正电的残基 Lys(675) 对 TRPV4 通道孔的性质没有明显影响。我们的研究结果描绘了 TRPV4 的孔隙区域，并首次深入了解其渗透途径的可能结构。
We have studied the molecular determinants of ion permeation through the TRPV4 channel (VRL-2, TRP12, VR-OAC, and OTRPC4). TRPV4 is characterized by both inward and outward rectification, voltage-dependent block by Ruthenium Red, a moderate selectivity for divalent versus monovalent cations, and an Eisenman IV permeability sequence. We identify two aspartate residues,Asp(672) and AsP682, as important determinants of the Ca2+ sensitivity of the TRPV4 pore. Neutralization of either aspartate to alanine caused a moderate reduction of the relative permeability for divalent cations and of the degree of outward rectification. Neutralizing both aspartates simultaneously caused a much stronger reduction of Ca2+ permeability and channel rectification and additionally altered the permeability order for monovalent cations toward Eisenman sequence II or I. Moreover, neutralizing Asp(682) but not AsP672 strongly reduces the affinity of the channel for Ruthenium Red. Mutations to Met(680), which is located at the center of a putative selectivity filter, strongly reduced whole cell current amplitude and impaired Ca2+ permeation. In contrast, neutralizing the only positively charged residue in the putative pore region, Lys(675), had no obvious effects on the properties of the TRPV4 channel pore. Our findings delineate the pore region of TRPV4 and give a first insight into the possible architecture of its permeation pathway.