期刊
EMBO JOURNAL
卷 28, 期 3, 页码 175-182出版社
NATURE PUBLISHING GROUP
DOI: 10.1038/emboj.2008.284
关键词
anion; antiporter; chloride; ion channel; proton transport
资金
- Telethon, Italy [GGP04018, GGP08064]
Several members of the CLC family are secondary active anion/proton exchangers, and not passive chloride channels. Among the exchangers, the endosomal ClC-5 protein that is mutated in Dent's disease shows an extreme outward rectification that precludes a precise determination of its transport stoichiometry from measurements of the reversal potential. We developed a novel imaging method to determine the absolute proton flux in Xenopus oocytes from the extracellular proton gradient. We determined a transport stoichiometry of 2 Cl-/1 H+. Nitrate uncoupled proton transport but mutating the highly conserved serine 168 to proline, as found in the plant NO3-/H+ antiporter atClCa, led to coupled NO3-/H+ exchange. Among several amino acids tested at position 168, S168P was unique in mediating highly coupled NO3-/H+ exchange. We further found that ClC-5 is strongly stimulated by intracellular protons in an allosteric manner with an apparent pK of similar to 7.2. A 2:1 stoichiometry appears to be a general property of CLC anion/proton exchangers. Serine 168 has an important function in determining anionic specificity of the exchange mechanism.
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