Identification of residues critical for Cu2+-mediated inhibition of glycine alpha1 receptors.
Identification of residues critical for Cu2+-mediated inhibition of glycine alpha1 receptors.
复制标题
鉴定对 Cu2 介导的甘氨酸 α1 受体抑制至关重要的残基。
DOI：
10.1016/j.neuropharm.2006.05.009
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发表时间：
2006
影响因子：
4.7
通讯作者：
Huang,Renqi
中科院分区：
文献类型：
--
作者：
Chen,Zhenglan;Dillon,GlennH;Huang,Renqi
内源二价阳离子Cu2+和Zn2+抑制甘氨酸受体的活性。虽然对锌离子对糖类的影响至关重要的残基已经确定，但对铜离子介导的抑制的决定因素知之甚少。在目前的研究中，我们评估了锌离子和铜离子介导的葡萄糖抑制的潜在共性。Cu2+对重组人甘氨酸α1受体有明显的抑制作用，IC50值为4.1±0.7μM。细胞外组氨酸残基的系统突变表明，突变的H2 15A大大减弱了Cu2+的抑制作用。用C取代H215产生的受体对Cu2+的敏感性与野生型相似。此外，用硫代特异性试剂[2-(三甲基氨乙基)乙基]甲基硫磺酸溴(MTSET)修饰H_215C，降低了H_215C受体对Cu2+的敏感性。然而，包括H107和H109在内的其他胞外组氨酸残基的突变并不影响Cu2+对甘氨酸电流的抑制。此外，锌离子抑制关键的两个苏氨酸残基(T112、T133)突变为丙氨酸对Cu2+诱导的抑制无影响(T133A)或仅有部分抑制作用(T112A)。T112A/H215A双突变对Cu2+介导的抑制的影响比单独突变更大。此外，低浓度的Cu2+可显著增强T112A/H215A突变受体的甘氨酸电流。我们的结果已经确定了Cu2+介导的葡萄糖抑制的关键决定因素。此外，我们首次证明了与锌离子介导的葡萄糖抑制相比，Cu2+介导的残基具有明显的差异。
Endogenous divalent cations Cu2+and Zn2+suppress the activity of glycine receptors (glyRs). Whereas residues critical for the effects of Zn2+on glyRs have been identified, little is known about the determinants of Cu2+-mediated inhibition. In the present studies, we have assessed the potential commonality of Zn2+and Cu2+-mediated inhibition of glyRs. Cu2+potently inhibited recombinant human glycine α1 receptors, with an IC50of 4.1±0.7μM. Systematic mutation of extracellular histidine residues revealed that mutation H215A greatly reduced the inhibitory modulation by Cu2+. Substitution of H215 with C produced receptors with Cu2+sensitivity similar to the wild type. Furthermore, modification of H215C with a thio-specific reagent, [2-(trimethylammonium)ethyl] methanethiosulfonate bromide (MTSET), reduced Cu2+sensitivity of H215C receptors. However, mutation of other extracellular histidine residues including H107 and H109, which are known inhibitory Zn2+coordination sites, failed to influence inhibition of glycine currents by Cu2+. Moreover, mutation to alanine of two threonine residues (T112, T133) critical for Zn2+inhibition had no effect (T133A) or only partial inhibitory effects (T112A) on Cu2+-induced inhibition. The double mutation, T112A/H215A, caused greater effects on Cu2+-mediated inhibition than either mutation alone. In addition, the glycine currents recorded from T112A/H215A mutant receptors were significantly potentiated by low concentrations of Cu2+. Our results have identified critical determinants of Cu2+-mediated inhibition of glyRs. Moreover, we demonstrate for the first time a clear difference in residues responsible for Cu2+-mediated compared to Zn2+-mediated inhibition of glyRs.