Functional complementation and genetic deletion studies of KirBac channels: activatory mutations highlight gating-sensitive domains.
Functional complementation and genetic deletion studies of KirBac channels: activatory mutations highlight gating-sensitive domains.
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DOI：
10.1074/jbc.m110.175687
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发表时间：
2010-12-24
期刊：
影响因子：
--
通讯作者：
Tucker SJ
中科院分区：
文献类型：
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作者：
Paynter JJ;Andres-Enguix I;Fowler PW;Tottey S;Cheng W;Enkvetchakul D;Bavro VN;Kusakabe Y;Sansom MS;Robinson NJ;Nichols CG;Tucker SJ
原核生物内向整流钾通道（KirBac）超家族与哺乳动物Kir通道同源。然而，对它们的调节或它们在体内的生理作用知之甚少。在这项研究中，我们使用了随机诱变和遗传互补K+-营养缺陷型大肠杆菌和酿酒酵母，以确定在一系列不同的KirBac通道的激活突变。我们还表明，KirBac6.1基因（slr 5078）是蓝藻集胞藻PCC 6803的正常生长所必需的。功能分析和分子动力学模拟选定的激活突变确定的区域内的幻灯片螺旋，跨膜螺旋，和C末端的KirBac通道活性的重要调节器的功能，以及区域接近KirBac3.1的选择性过滤器，可能有影响门控。特别是，在TM 2中鉴定的突变有利于KirBac通道门控的模型，其中在螺旋束交叉处的孔的打开起着比最近提出的重要得多的作用。
The superfamily of prokaryotic inwardly rectifying (KirBac) potassium channels is homologous to mammalian Kir channels. However, relatively little is known about their regulation or about their physiological role in vivo. In this study, we have used random mutagenesis and genetic complementation in K+-auxotrophic Escherichia coli and Saccharomyces cerevisiae to identify activatory mutations in a range of different KirBac channels. We also show that the KirBac6.1 gene (slr5078) is necessary for normal growth of the cyanobacterium Synechocystis PCC6803. Functional analysis and molecular dynamics simulations of selected activatory mutations identified regions within the slide helix, transmembrane helices, and C terminus that function as important regulators of KirBac channel activity, as well as a region close to the selectivity filter of KirBac3.1 that may have an effect on gating. In particular, the mutations identified in TM2 favor a model of KirBac channel gating in which opening of the pore at the helix-bundle crossing plays a far more important role than has recently been proposed.