Laura Lagostena1, Giovanni Zifarelli2, Alessandra Picollo3. 1. Dulbecco Telethon Laboratory, Istituto di Biofisica, Consiglio Nazionale delle Ricerche, Genova, Italy; and. 2. Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, UK giovanni.zifarelli@dpag.ox.ac.uk apicollo@dti.telethon.it. 3. Dulbecco Telethon Laboratory, Istituto di Biofisica, Consiglio Nazionale delle Ricerche, Genova, Italy; and giovanni.zifarelli@dpag.ox.ac.uk apicollo@dti.telethon.it.
Abstract
BACKGROUND: The mechanism of anion selectivity in the human kidney chloride channels ClC-Ka and ClC-Kb is unknown. However, it has been thought to be very similar to that of other channels and antiporters of the CLC protein family, and to rely on anions interacting with a conserved Ser residue (Sercen) at the center of three anion binding sites in the permeation pathway Scen. In both CLC channels and antiporters, mutations of Sercen alter the anion selectivity. Structurally, the side chain of Sercen of CLC channels and antiporters typically projects into the pore and coordinates the anion bound at Scen. METHODS: To investigate the role of several residues in anion selectivity of ClC-Ka, we created mutations that resulted in amino acid substitutions in these residues. We also used electrophysiologic techniques to assess the properties of the mutants. RESULTS: Mutations in ClC-Ka that change Sercen to Gly, Pro, or Thr have only minor effects on anion selectivity, whereas the mutations in residues Y425A, F519A, and Y520A increase the NO3 -/Cl- permeability ratio, with Y425A having a particularly strong effect. CONCLUSION: s ClC-Ka's mechanism of anion selectivity is largely independent of Sercen, and it is therefore unique in the CLC protein family. We identified the residue Y425 in ClC-Ka-and the corresponding residue (A417) in the chloride channel ClC-0-as residues that contribute to NO3 - discrimination in these channels. This work provides important and timely insight into the relationship between structure and function for the kidney chloride channels ClC-Ka and ClC-Kb, and for CLC proteins in general.
BACKGROUND: The mechanism of anion selectivity in the human kidney chloride channels ClC-Ka and ClC-Kb is unknown. However, it has been thought to be very similar to that of other channels and antiporters of the CLC protein family, and to rely on anions interacting with a conserved Ser residue (Sercen) at the center of three anion binding sites in the permeation pathway Scen. In both CLC channels and antiporters, mutations of Sercen alter the anion selectivity. Structurally, the side chain of Sercen of CLC channels and antiporters typically projects into the pore and coordinates the anion bound at Scen. METHODS: To investigate the role of several residues in anion selectivity of ClC-Ka, we created mutations that resulted in amino acid substitutions in these residues. We also used electrophysiologic techniques to assess the properties of the mutants. RESULTS: Mutations in ClC-Ka that change Sercen to Gly, Pro, or Thr have only minor effects on anion selectivity, whereas the mutations in residues Y425A, F519A, and Y520A increase the NO3 -/Cl- permeability ratio, with Y425A having a particularly strong effect. CONCLUSION: s ClC-Ka's mechanism of anion selectivity is largely independent of Sercen, and it is therefore unique in the CLC protein family. We identified the residue Y425 in ClC-Ka-and the corresponding residue (A417) in the chloride channel ClC-0-as residues that contribute to NO3 - discrimination in these channels. This work provides important and timely insight into the relationship between structure and function for the kidney chloride channels ClC-Ka and ClC-Kb, and for CLC proteins in general.
Authors: Karl P Schlingmann; Martin Konrad; Nikola Jeck; Petra Waldegger; Stephan C Reinalter; Martin Holder; Hannsjörg W Seyberth; Siegfried Waldegger Journal: N Engl J Med Date: 2004-03-25 Impact factor: 91.245
Authors: R Birkenhäger; E Otto; M J Schürmann; M Vollmer; E M Ruf; I Maier-Lutz; F Beekmann; A Fekete; H Omran; D Feldmann; D V Milford; N Jeck; M Konrad; D Landau; N V Knoers; C Antignac; R Sudbrak; A Kispert; F Hildebrandt Journal: Nat Genet Date: 2001-11 Impact factor: 38.330
Authors: Anselm A Zdebik; Giovanni Zifarelli; Eun-Yeong Bergsdorf; Paolo Soliani; Olaf Scheel; Thomas J Jentsch; Michael Pusch Journal: J Biol Chem Date: 2007-12-06 Impact factor: 5.157