The KCNQ1 voltage-gated potassium channel regulates the repolarization of cardiac cells, and a plurality of point mutations in its voltage-sensing domain (VSD) are associated with toxic gain or loss of pore function, resulting in disease. As is the case with many disease-associated membrane proteins, there are hundreds of human variants of interest identified for KCNQ1; however, a significant portion of these variants have not been characterized in relation to their functional and disease associations. Additionally, as the VSD consists of four transmembrane helices, studies into dynamic structural differences among KCNQ1 VSD variants are hindered by the current limitations and deficits in the high-resolution structure determination of membrane proteins. Here, we use native ion mobility-mass spectrometry and collision-induced unfolding (CIU) to address the need for a high throughput-compatible method for the structural characterization of membrane protein variants of unknown significance using the KCNQ1 VSD as a model system. We perform CIU on wild-type and three mutant KCNQ1 VSD forms associated with the toxic gain or loss of function and show through both automated feature detection and comprehensive difference analysis of the CIU data sets that the variants are clearly grouped by function and disease association. We also construct a classification scheme based on the CIU data sets, which is able to differentiate the variant functional groups and classify a recently characterized variant to its correct grouping. Further, we probe the stability of the KCNQ1 VSD variants when liberated from C12E8 micelles at pH 8.0 and find preliminary evidence that the R231C mutation associated with the gain of the pore function is destabilized relative to the wild-type and loss of function variants.

中文翻译：

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碰撞引发的展开区分 KCNQ1 电压传感器域的功能变体。

KCNQ1电压门控钾通道调节心脏细胞的复极化，其电压感应域（VSD）中的多个点突变与毒性增加或孔功能丧失相关，从而导致疾病。与许多与疾病相关的膜蛋白一样，KCNQ1 已鉴定出数百种人类感兴趣的变体。然而，这些变体中有很大一部分尚未就其功能和疾病关联进行表征。此外，由于 VSD 由四个跨膜螺旋组成，因此对 KCNQ1 VSD 变体之间动态结构差异的研究受到目前膜蛋白高分辨率结构测定的局限性和缺陷的阻碍。这里，我们使用天然离子淌度质谱法和碰撞诱导展开 (CIU) 来满足使用 KCNQ1 VSD 作为模型系统对具有未知意义的膜蛋白变体进行结构表征的高通量兼容方法的需求。我们对与毒性获得或丧失功能相关的野生型和三种突变型 KCNQ1 VSD 形式进行 CIU，并通过对 CIU 数据集的自动特征检测和综合差异分析表明，变体按功能和疾病关联明确分组。我们还构建了基于 CIU 数据集的分类方案，该方案能够区分变体功能组并将最近表征的变体分类为其正确分组。更远，