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XB-ART-39180
EMBO J 2009 Feb 04;283:175-82. doi: 10.1038/emboj.2008.284.
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Conversion of the 2 Cl(-)/1 H+ antiporter ClC-5 in a NO3(-)/H+ antiporter by a single point mutation.

Zifarelli G , Pusch M .


Abstract
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 approximately 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.

PubMed ID: 19131966
PMC ID: PMC2637338
Article link: EMBO J
Grant support: [+]

Species referenced: Xenopus
Genes referenced: clcn5 nbl1

References [+] :
Accardi, Secondary active transport mediated by a prokaryotic homologue of ClC Cl- channels. 2004, Pubmed