Magnesium Channel Cation Selectivity

镁通道阳离子选择性

基本信息

批准号：
8667478
负责人：
MICHAEL E MAGUIRE
金额：
$ 29.83万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-24 至 2016-05-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Mg2+ is an essential element. The basis of Mg2+ selectivity, binding and coordination with enzymes and small molecules is well understood but the molecular basis by which transport systems achieve Mg2+ selectivity over other cations is not understood in any system. The structures of the CorA and MgtE Mg2+ channels demonstrate clearly that channel selectivity for Mg2+ cannot be explained by analogy to the chemistry of selective Mg2+ sites in any enzyme system. The CorA Mg2+ channel is the primary source of Mg2+ for ~50% of all Bacteria and Archaea while MgtE is the primary source for the other half of each kingdom. Both have multiple homologs in eukaryotes, including humans. Electrophysiological data show that both CorA and MgtE are true ion channel with surprisingly high conductance, >100 pS, a flux rate >107 Mg2+ ions/sec. This very high rate poses serious mechanistic issues. The rate of dehydration of Mg2+ is only 105/sec, 100-1000-fold slower than the flux rate. Uniquely among all known ion channels, both CorA and MgtE initially bind a fully hydrated Mg2+ ion, which must contribute to the channel's selectivity for Mg2+. To mediate flux of a largely dehydrated cation, the channel must in some manner accelerate the rate of dehydration. However, electrostatic interactions are not involved in this process or in ion flux. The study of these two ion channels, which have quite different structures, provides an opportunity to dissect the distinct chemistry involved in Mg2+ selectivity. Aim 1 will investigate the mechanism of selectivity of CorA for Mg2+ through i) electrophysiological analysis of site-directed mutants expressed in giant cells or reconstituted in lipid, ii) hydrogen-deuterium exchange primarily of loop and pore residues, and iii) X-ray crystallographic approaches using cations with very slow rates of dehydration and cysteine mutations in transmembrane segment 2 of CorA. The latter generate crosslinked oligomers which appear to be trapped in a conformation different from the closed state, previously solved. Their crystallization may provide additional conformational information for CorA. Aim 2 will explore analogous issues for MgtE using the same approaches.

描述（由申请人提供）：MG2+是必不可少的元素。人们充分了解了MG2+选择性，与酶和小分子的结合和协调的基础，但是在任何系统中尚不理解运输系统实现MG2+选择性的分子基础。 CORA和MGTE MG2+通道的结构清楚地证明了MG2+的通道选择性不能与任何酶系统中选择性MG2+位点的化学相似。 CORA MG2+通道是所有细菌和古细菌的50％的MG2+的主要来源，而MGTE是每个王国另一半的主要来源。两者在包括人类在内的真核生物中都有多种同源物。电生理数据表明，CORA和MGTE都是真正的离子通道，其电导率高于100 ps，通量速率> 107 mg2+离子/秒。这种很高的比率提出了严重的机械问题。 MG2+的脱水速率仅为105/sec，比通量速率慢100-1000倍。在所有已知的离子通道中，Cora和MGTE都独特地结合了完全水合的Mg2+离子，这必须有助于该通道对MG2+的选择性。为了介导很大程度上脱水的阳离子的通量，该通道必须以某种方式加速脱水速率。但是，静电相互作用不参与此过程或离子通量。对具有完全不同结构的这两个离子通道的研究提供了一个机会，可以剖析与MG2+选择性相关的不同化学反应。 Aim 1 will investigate the mechanism of selectivity of CorA for Mg2+ through i) electrophysiological analysis of site-directed mutants expressed in giant cells or reconstituted in lipid, ii) hydrogen-deuterium exchange primarily of loop and pore residues, and iii) X-ray crystallographic approaches using cations with very slow rates of dehydration and cysteine mutations in transmembrane segment 2 of CorA.后者产生的交联寡聚物似乎被困在与以前解决的封闭状态不同的构象中。它们的结晶可以为Cora提供其他构象信息。 AIM 2将使用相同的方法探索MGTE的类似问题。