Department of Biochemistry, Howard Hughes Medical Institute, Brandeis University, Waltham, MA 02454, USA.
Proc Natl Acad Sci U S A. 2012 Sep 18;109(38):15289-94. doi: 10.1073/pnas.1210896109. Epub 2012 Sep 4.
A subclass of bacterial CLC anion-transporting proteins, phylogenetically distant from long-studied CLCs, was recently shown to be specifically up-regulated by F(-). We establish here that a set of randomly selected representatives from this "CLC(F)" clade protect Escherichia coli from F(-) toxicity, and that the purified proteins catalyze transport of F(-) in liposomes. Sequence alignments and membrane transport experiments using (19)F NMR, osmotic response assays, and planar lipid bilayer recordings reveal four mechanistic traits that set CLC(F) proteins apart from all other known CLCs. First, CLC(F)s lack conserved residues that form the anion binding site in canonical CLCs. Second, CLC(F)s exhibit high anion selectivity for F(-) over Cl(-). Third, at a residue thought to distinguish CLC channels and transporters, CLC(F)s bear a channel-like valine rather than a transporter-like glutamate, and yet are F(-)/H(+) antiporters. Finally, F(-)/H(+) exchange occurs with 1:1 stoichiometry, in contrast to the usual value of 2:1.
最近发现，细菌 CLC 阴离子转运蛋白的一个亚类与长期研究的 CLC 亲缘关系较远，其特异性受 F(-)调控。本文中我们确定了该“CLC(F)”分支中一组随机挑选的代表蛋白可保护大肠杆菌免受 F(-)毒性的影响，并且纯化的蛋白可在脂质体中催化 F(-)的转运。序列比对和使用 (19)F NMR、渗透响应测定和平面脂质双层记录的膜转运实验揭示了将 CLC(F)蛋白与所有其他已知 CLC 区分开来的四个机制特征。首先，CLC(F)缺乏形成经典 CLC 中阴离子结合位点的保守残基。其次，CLC(F)对 F(-)表现出高于 Cl(-)的高阴离子选择性。第三，在一个被认为区分 CLC 通道和转运蛋白的残基处，CLC(F)具有类似通道的缬氨酸而不是类似转运蛋白的谷氨酸，但却是 F(-)/H(+)反向转运体。最后，F(-)/H(+)交换以 1:1 的化学计量发生，与通常的 2:1 值相反。
