title: Basis of substrate binding and conservation of selectivity in the CLC family of channels and transporters.
abstract: Ion binding to secondary active transporters triggers a cascade of conformational rearrangements resulting in substrate translocation across cellular membranes. Despite the fundamental role of this step, direct measurements of binding to transporters are rare. We investigated ion binding and selectivity in CLC-ec1, a H(+)-Cl(-) exchanger of the CLC family of channels and transporters. Cl(-) affinity depends on the conformation of the protein: it is highest with the extracellular gate removed and weakens as the transporter adopts the occluded configuration and with the intracellular gate removed. The central ion-binding site determines selectivity in CLC transporters and channels. A serine-to-proline substitution at this site confers NO(3)(-) selectivity upon the Cl(-)-specific CLC-ec1 transporter and CLC-0 channel. We propose that CLC-ec1 operates through an affinity-switch mechanism and that the bases of substrate specificity are conserved in the CLC channels and transporters.
authors.affiliation: Department of Molecular Physiology and Biophysics, Roy J. and Lucille A. Carver College of Medicine, University of Iowa, Iowa City, Iowa, USA.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Picollo
authors.forename: Mattia
authors.initials: M
authors.lastname: Malvezzi
authors.forename: Jon C D
authors.initials: JC
authors.lastname: Houtman
authors.forename: Alessio
authors.initials: A
authors.lastname: Accardi
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D000838:Anions / Q000378:metabolism*; D001665:Binding Sites; D018118:Chloride Channels / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002713:Chlorine / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009566:Nitrates / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity
chemical_list: D000838:Anions; D018118:Chloride Channels; C508370:ClC protein, E coli; D029968:Escherichia coli Proteins; D009566:Nitrates; D002713:Chlorine