Insights into the structural basis for zinc inhibition of the glycine receptor
Insights into the structural basis for zinc inhibition of the glycine receptor
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DOI：
10.1074/jbc.m300097200
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发表时间：
2003-08-01
影响因子：
4.8
通讯作者：
Lynch, JW
中科院分区：
文献类型：
--
作者：
Nevin, ST;Cromer, BA;Lynch, JW
甘氨酸受体(GlyR)α(1)亚单位中的组氨酸107和109是锌结合抑制位点的决定因素。根据与乙酰胆碱结合蛋白晶体结构的同源性对GlyRα(1)亚基胞外结构域的建模，我们假设抑制性锌结合在亚基界面的前庭管腔内，在那里它被来自一个亚基的His(107)和来自相邻亚基的His(109)连接。这是通过共表达包含H107A突变的阿尔法(1)亚基和含有H109A突变的阿尔法(1)亚基来检验的。尽管当任何一个突变单独整合到所有五个亚基中时，对锌抑制的敏感性显著降低，但由H107A突变亚基和H109A突变亚基共表达形成的GlyR显示出与野生型α(1)同源GlyR相似的锌抑制敏感性。这有力地证明了抑制性锌在相邻的α(1)亚基之间的界面上是协调的。没有发现β亚基参与抑制锌的协调的证据，这表明每个α(1)β受体最多有两个锌结合位点足以最大限度地抑制锌。我们的数据还表明，两个锌结合位点足以显著抑制α(1)异构体。锌在相邻α(1)亚基界面的结合可以限制亚基间的运动，为锌抑制通道激活提供了一种可行的机制。
Histidines 107 and 109 in the glycine receptor ( GlyR) alpha(1) subunit have previously been identified as determinants of the inhibitory zinc-binding site. Based on modeling of the GlyR alpha(1) subunit extracellular domain by homology to the acetylcholine-binding protein crystal structure, we hypothesized that inhibitory zinc is bound within the vestibule lumen at subunit interfaces, where it is ligated by His(107) from one subunit and His(109) from an adjacent subunit. This was tested by co-expressing alpha(1) subunits containing the H107A mutation with alpha(1) subunits containing the H109A mutation. Although sensitivity to zinc inhibition is markedly reduced when either mutation is individually incorporated into all five subunits, the GlyRs formed by the co-expression of H107A mutant subunits with H109A mutant subunits exhibited an inhibitory zinc sensitivity similar to that of the wild type alpha(1) homomeric GlyR. This constitutes strong evidence that inhibitory zinc is coordinated at the interface between adjacent alpha(1) subunits. No evidence was found for beta subunit involvement in the coordination of inhibitory zinc, indicating that a maximum of two zinc-binding sites per alpha(1)beta receptor is sufficient for maximal zinc inhibition. Our data also show that two zinc-binding sites are sufficient for significant inhibition of alpha(1) homomers. The binding of zinc at the interface between adjacent alpha(1) subunits could restrict intersubunit movements, providing a feasible mechanism for the inhibition of channel activation by zinc.