The yeast Trk1 polypeptide, like other members of the Superfamily of K Transporters (SKT proteins) consists of four Membrane-Pore-Membrane motifs (MPMs A-D) each of which is homologous to a single K-channel subunit. SKT proteins are thought to have evolved from ancestral K-channels via two gene duplications and thus single MPMs might be able to assemble when located on different polypeptides. To test this hypothesis experimentally we generated a set of partial gene deletions to create alleles encoding one, two, or three MPMs, and analysed the cellular localisation and interactions of these Trk1 fragments using GFP tags and Bimolecular Fluorescence Complementation (BiFC). The function of these partial Trk1 proteins either alone or in combinations was assessed by expressing the encoding genes in a K⁺-uptake deficient strain lacking also the K-channel Tok1 (trk1,trk2,tok1Δ) and (i) analysing their ability to promote growth in low [K⁺] media and (ii) by ion flux measurements using “microelectrode based ion flux estimation” (MIFE). We found that proteins containing only one or two MPM motifs can interact with each other and assemble with a polypeptide consisting of the rest of the Trk system to form a functional K⁺-translocation system.

与K转运蛋白超家族（SKT蛋白）的其他成员一样，酵母Trk1多肽由四个膜-孔-膜基序（MPMs AD）组成，每个基序与单个K通道亚基同源。人们认为SKT蛋白是通过两个基因复制从祖先K通道进化而来的，因此单个MPM在位于不同多肽上时可能能够组装。为了通过实验测试此假设，我们生成了一组部分基因缺失，以创建编码一个，两个或三个MPM的等位基因，并使用GFP标签和双分子荧光互补（BiFC）分析了这些Trk1片段的细胞定位和相互作用。通过在K ^+中表达编码基因来评估这些部分Trk1蛋白单独或组合的功能。缺乏K通道Tok1（trk1，trk2，tok1Δ）的吸收不足菌株，以及（i）分析其在低[K ⁺ ]培养基中促进生长的能力，以及（ii）使用“基于微电极的离子通量估算”进行离子通量测量”（MIFE）。我们发现，仅包含一个或两个MPM基序的蛋白质可以彼此相互作用，并与由Trk系统其余部分组成的多肽组装在一起，形成功能性K ^+-转运系统。